Registration Dossier

Administrative data

Key value for chemical safety assessment

Effects on fertility

Description of key information

No adverse effects were observed in male and female Sprague-Dawley rats in a 54 days combined repeated dose toxicity study with the reproduction/developmental toxicity screening test conducted according to OECD 422 (BASF, 2007). Animals were exposed orally (gavage) to 0, 100, 300 and 1000 mg/kg bw/day. Under the conditions of this test the NOAEL for reproductive performance and fertility is 1000 mg/kg bw/day for F0 parental rats.

Link to relevant study records
Reference
Endpoint:
screening for reproductive / developmental toxicity
Type of information:
experimental study
Adequacy of study:
key study
Study period:
31 January 2006 - 28 September 2006
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
guideline study with acceptable restrictions
Remarks:
Well documented GLP study performed according to OECD Guideline 422 and EPA/OPPTS 870.3650, on the hydrochloride salt of the submission substance. Erroneously, several females of all test groups including the controls were gavaged with application volumes being 0.1 -0.3 mL above or below the target volumes on two days. These marginal and transient deviations from the study protocol are not considered to have any influence on the outcome of this study.
Qualifier:
according to
Guideline:
OECD Guideline 422 (Combined Repeated Dose Toxicity Study with the Reproduction / Developmental Toxicity Screening Test)
Deviations:
yes
Remarks:
dosing volumes (see field 'Any other information on materials and methods incl. tables)
Qualifier:
according to
Guideline:
other: EPA/OPPTS 870.3650: Combined Repeated Dose Toxicity Study With the Reproduction/Developmental Toxicity Screening Test (July 2000)
Deviations:
yes
Remarks:
dosing volumes (see field 'Any other information on materials and methods incl. tables)
GLP compliance:
yes (incl. certificate)
Limit test:
no
Specific details on test material used for the study:
- Name of test material (as cited in study report): Hydrochloride salt of 3-Methoxypropylamine
- Molecular formula (if other than submission substance): C4 H12 Cl N O
- Molecular weight (if other than submission substance): 125.6 g/mole
- Substance type: Aqueous solution / liquid, orange, clear
- Physical state: Aqueous solution/liquid
- Lot/batch No.: BA 1464
- Storage conditions: Room temperature
- Other:
Test substance No.: 05/0675-1
Concentration of stock solution: 67.2 g 3-Methoxypropylamine hydrochloride per 100 g aqueous solution
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
- Strain: Crl:WI(Han)
- Age at study initiation: 11 - 13 weeks
- Weight at study initiation: male animals (264.0 g) and female animals (200.2 g)
- Fasting period before study: food and water were available ad libitum except during the fasting period (16 to 20 hours prior to necropsy) and measurement of motor activity)
- Housing: The rats were housed individually in type DK III stainless steel wire mesh cages supplied by Becker & Co., Castrop-Rauxel, Germany (floor area of about 800 cm2), with following exceptions: for the overnight mating the females were put into the cages of the males; from day 18 p.c. until sacrifice, the pregnant animals and their litters were housed in Makrolon type M III cages (floor area about 800 cm2). The M III cages were also supplied by Becker & Co. Pregnant females were provided with nesting material (cellulose wadding) toward the end of pregnancy. The cages with the test animals were arranged on the racks in such way that uniform experimental conditions (ventilation and light) were ensured. Motor activity measurements were conducted in Polycarbonate cages with wire covers from Ehret, Emmendingen, Germany (floor area about 800 cm2) and small amounts of absorbant material.
- Diet (e.g. ad libitum): The food used was ground Kliba maintenance diet mouse/rat "GLP", supplied by Provimi Kliba SA, Kaiseraugst, Switzerland.
- Water (e.g. ad libitum): from water bottles available ad libitum (except during the fasting period and measurement of motor activity).
- Acclimation period: about 6 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 20-24°C
- Humidity (%): 30-70%
- Photoperiod (hrs dark / hrs light): 12 hours light, 12 hours dark
Route of administration:
oral: gavage
Vehicle:
water
Details on exposure:
PREPARATION OF DOSING SOLUTIONS:
For preparation of the administration solutions, the test substance was weighed in a graduated measuring flask (corresponding to the dose group), topped up with tap water and dissolved by shaking.

The test substance solutions were prepared at the beginning of the administration period and thereafter at intervals that took into account the stability of the test substance preparation.

The pH-value of each test substance preparation was measured before dosing. All dosing solutions were in the pH range of 6.0 to 7.5.
Details on mating procedure:
Each of the male and female animals was mated overnight in a 1:1 ratio for a maximum of 2 weeks. Throughout the mating period, each female animal was paired with a predetermined male animal from the same dose group. The animals were paired by placing the female in the cage of the male mating partner about 4.00 p.m. until 7.00-9.00 a.m. of the following morning. Deviations from the specified times were possible on weekends and public holidays and were reported in the raw data. a vaginal smear was prepared after each mating and examined for sperm. If sperm was detected, pairing of the animals was discontinued. The day on which sperm was detected was denoted "day 0" and the following day "day 1" post coitum.
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
The analytical studies were carried out in compliance with GLP.
Method: Potentiometric titration
Discussion of results: Assuming a content of the test item of 100% at 0h, at least approximately 94% of the test item were stable in the vehicle for at least 11 days at ambient temperature.
Duration of treatment / exposure:
The duration of treatment covered a 2 week premating period and mating period in both sexes, approximately 1 week post-mating in males, and the entire gestation period and 4 days of lactation in females.
Frequency of treatment:
Daily at the same time in the morning (exception: no administration to animals being in labor).
Dose / conc.:
0 mg/kg bw/day (nominal)
Dose / conc.:
100 mg/kg bw/day (nominal)
Dose / conc.:
300 mg/kg bw/day (nominal)
Dose / conc.:
1 000 mg/kg bw/day (nominal)
No. of animals per sex per dose:
12
Control animals:
yes, concurrent vehicle
Details on study design:
The oral route was selected as it is considered to be the route whereby the majority of exposure would occur.
Positive control:
not applicable
Parental animals: Observations and examinations:
CAGE SIDE OBSERVATIONS: Yes
- At least once daily for any signs of morbidity, pertinent behavioral changes and signs of overt toxicity. Abnormalities and changes were documented for each animal.

DETAILED CLINICAL OBSERVATIONS: Yes
- 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 sides of 25 cm high). The following parameters were examined: abnormal behavior during "handling", fur, skin, salivation, nose discharge, lacrimation, pupil size, posture, palpebral closure, respiration, tremors, convulsions, abnormal movements, impairment of gait, activity/arousal level, feces (appearance/consistency), urine, other findings

BODY WEIGHT: Yes
- In general, the body weight of the male and female parental animals was determined once a week at the same time of the day (in the morning).
The body weight change of the animals was calculated from these results.
The following exceptions are notable for the female animals:
- During the mating period the parental females were weighed on the day of positive evidence of sperm (day 0 post coitum) and on days 7, 14 and 20 post coitum.
- Females with litter were weighed on the day of parturition (day 0 post partum) and on day 4 post partum.
- Females without a litter were weighed weekly. These body weight data were solely used for the calculations of the dose volume.

FOOD CONSUMPTION: Yes
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
- Food consumption of the F0 females with evidence of sperm was determined on days 0, 7, 14 and 20 post coitum
- Food consumption of F0 females, which gave birth to a litter was determined on days 0 and 4 post partum.
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: Determination of implantation sites: After sacrifice of the female animals the uterus and ovaries were removed and the implantation sites were counted. To determine the number of implantation sites in apparently non-pregnant animals, the uteri from those females were stained in 10% ammonium sulfide solution for about 5 minutes according to the method of SALEWSKI (1964). Then, the respective uteri were rinsed carefully with fresh tap water. The implantation sites were recorded for the calculation of the postimplantation loss.
Oestrous cyclicity (parental animals):
No data
Sperm parameters (parental animals):
No data
Litter observations:
Pup numbers and status at delivery:
all pups delivered from the F0 partents were examined as soon as possible on the day of birth to determine the total number of pups and the number of liveborn and stillborn pups in each litter. Pups, which died before the first determination of their status on the day of birth, were defined as stillborn pups.
Pup viability/mortality:
In general a check was made for any dead or moribund pups twice daily on workdays (once in the morning and onec in the afternoon) or as a rule, only in the morning on Saturdays, Sundays or public holidays.
The number and percentage of dead pups on the day of birth (day 0 post partum) and pups dying between days 1-4 of the lactation period were determined; however, pups which died accidentally and pups which were sacrificed due to maternal death were not included in these calculations. The number of live pups/litter was calculated on the day of birth, and on lactation day 4. Furthermore the viability index was calculated according to the following formula:
Viability index (%) = (number of live pups on day 4 after birth/number of liveborn pups on the day of birth) x 100
Sex ratio:
On the day of birth (day 0) the sex of the pups was determined by observing the distance between the anus and the base of the genital tubercle; normally, the anogenital distance is considerably greater in male than in female pups. The sex of the pups finally confirmed at necropsy.
The sex ratio was calculated at day 0 and day 4 after birth according to the following formula:
Sex ratio = (number of live male or female pups on day 0/4 / number of live male and female pups on day 0/4) x 100
Pup clinical observations
The live pups were examined daily for clinical symptoms (including gross-morphological findings) during the clinical inspection of the dams. If pups showed particular findings, these were documented with the dam concerned.
Pup body weight data:
The pups were weighed one day after birth (day 1 post partum) and on day 4 after birth.
Pups' body weight change was calculated from these results.
Furthermore the body weights on day 1 post partum were used for the calculation of "runts" (pups, which weighed more than 25% less than the mean weight of the respective control pups).
The individual weights were always determined at about the same time of the day (in the morning).

Postmortem examinations (parental animals):
Necropsy: The animals were sacrificed by decapitation under CO2 anesthesia. The exsanguinated animals were necropsied and assessed by gross pathology.
Organ weights: the weights of the following organs were determined in all animals sacrificed at scheduled dates: anesthetized animals, liver, kidneys, adrenal glands, testes, epididymides, seminal vesicle, prostate, ovaries, uterus, thymus, spleen, brain, heart
Organ/Tissue fixation: the following organs or tissues were fixed in 4% formaldehyde solution: all gross lesions, brain, spinal cord (cervical, thoracic and lumbar cord), sciatic nerve, pituitary gland, salivary glands, thyroid glands, parathyroid glands, adrenal glands, prostate gland, seminal vesicles, coagulation glands, uterus, oviducts, vagina, female mammary gland, thymus, lymph nodes (axillar and mesenteric), spleen, trachea, lungs, heart, aorta, liver, pancreas, kidneys, esophagus, stomach (forestomach and glandular stomach), duodenum, jejunum, ileum, cecum, colon, rectum, urinary bladder, sternum with marrow, bone marrow (femur), eyes with optic nerve, femur with knee joint, skin and skeletal muscle.
Histopathology: all gross lesions, trachea, lungs, liver, kidneys, spleen, adrenal glands, heart, brain, spinal cord (cervical, thoracic, lumbar), sciatic nerve, thyroid glands/parathyroid glands, testes, epididymides, ovaries, oviducts, uterus, vagina, prostate glands, seminal vesicles, coagulation glands, thymus, lymph nodes, stomach, duodenum, jejunum, ileum, cecum, colon, rectum, urinary bladder, bone marrow.
Postmortem examinations (offspring):
All surviving pups were sacrificed (by means of CO2) on day 4 p.p., were examined externally, eviscerated and their organs were assessed macroscopically. Stillborn pups and all pups died before schedule, were examined externally, eviscerated and their organs were assessed macroscopically. All pups were discarded after their evaluation.
Statistics:
Food consumption, body weight and body weight change, number of mating days, duration of gestation, number of pups delivered per litter, implantation sites, post implantation loss: Dunnet's test (two-sided) for the hypothesis of equal means)
Male and female mating index, male and female fertility index, gestation index, females with liveborn pups, 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: Pairwise comparison of each dose group with the control group using Fisher's exact test for the hypothesis of equal proportions
Proportions of affected pups per litter with necropsy observations: pairwise comparison of each dose group with the control group using the Wilcoxon-test (on-sided) for the hypothesis of equal medians
Feces, rearing, grip strength of forelimbs and hindlimbs, landing footsplay test, motor activity: non-parametric one-way analysis using Kruskal-Wallis test (two-sided). If the resulting p-value was equal or less than 0.05, a pairwise comparison of each dose group with the control group was performed using Wilcoxon-test (two-sided) for the equal medians.
Clinical pathology parameters, except reticulocytes and differential blood count: non-parametric one-way analysis using Kruskal-Wallis test (two-sided). If the resulting p-value was equal or less than 0.05, a pairwise comparison of each dose group with the control group was performed using wilcoxon-test (two-sided) for the equal medians.
Urinalysis, except volume, color, turbidity and specific gravity: pairwise comparison of each dose group with the control group using Fisher's exact test for the hypothesis of equal proportions
Weight of anesthesized animals and absolute and relative organ weights: Kruskal-Wallis and Wilcoxon test
Reproductive indices:
The pairing partners, the number of mating days until vaginal sperm was detected in female animals, and the gestational status of the females were recorded for the F0 breeding pairs. For the males, mating and fertility indices were calculated according to the following formulas:
Male mating index (%) = (number of males with confirmed mating*/number of males placed with females) x 100
*: 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) x 100
*: defined by a female with implants in utero

Female mating index (%) = (number of females mated*/number of females placed with males) x 100
*: 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 **) x 100
*: 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*) x 100
*: defined as the number of females with implants in utero

Offspring viability indices:
The total number of pups delivered and the number of liveborn and stillborn pups were noted and the live birth index was calculated according to the following formulas:
Live birth index (%) = (number of liveborn pups at birth/total number of pups born) x 100
Moreover, after sacrifice of the female animals, the implantation sites were counted and the post implantation loss was calculated for each individual pregnant animal according to the following formula:
Post implantation loss (%) = (number of implantations - number of pups delivered/number of implantations) x 100
Clinical signs:
no effects observed
Description (incidence and severity):
Clinical observations for signs of general toxicity revealed post-dose salivation in 5 high dose males and all high dose females. Moreover, urine discoloration was observed for all mid and high dose rats. Both findings are considered to be without toxicological relevance and are, if seen in isolation, not assessed as being adverse.
Dermal irritation (if dermal study):
not examined
Mortality:
no mortality observed
Body weight and weight changes:
no effects observed
Food consumption and compound intake (if feeding study):
no effects observed
Food efficiency:
not examined
Water consumption and compound intake (if drinking water study):
not examined
Ophthalmological findings:
not examined
Haematological findings:
no effects observed
Clinical biochemistry findings:
effects observed, treatment-related
Urinalysis findings:
effects observed, treatment-related
Description (incidence and severity):
Reduced urine volumes with subsequently increased specific gravity were excreted by female rats of the top dose group (1000 mg/kg body weight/day). It is likely, that this was caused by a decreased water intake. It is concluded that the changes in urinalysis are not caused by a direct toxic effect of the test compound on the kidney and are not considered adverse in nature. This assessment is supported by the fact, that kidney weights as well as gross and histopathological evaluations of this organ did not give any indications for substance-induced alterations in the high dose females.
Behaviour (functional findings):
no effects observed
Organ weight findings including organ / body weight ratios:
no effects observed
Gross pathological findings:
no effects observed
Description (incidence and severity):
Clinical pathology examinations suggested a slightly impaired liver function of the high dose females. The production of albumin and some other proteins, which are synthesized in the liver cells, was probably decreased. Additionally, the ability of the hepatocytes to eliminate bilirubin from the body by conjugation was probably diminished. Moreover, higher triglyceride levels were found in the blood of these dams. The signs indicating a disturbed liver function are considered as adverse substance-induced effects. However, this original finding is challenged. Taking into account all the other information available on this study (no histopathological correlation as liver observations are similar to the controls), these changes seem the result of an adaptive process rather than an adverse effect. This assumption is also supported by the fact that no changes have been observed in ALT, AST, ALP and GCP levels.
Neuropathological findings:
not examined
Histopathological findings: non-neoplastic:
no effects observed
Other effects:
no effects observed
Reproductive function: oestrous cycle:
not examined
Reproductive function: sperm measures:
not examined
Reproductive performance:
no effects observed
Key result
Dose descriptor:
NOAEL
Remarks:
reproductive performance and fertility
Effect level:
1 000 mg/kg bw/day (actual dose received)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
other: No adverse effects at 1000 mg/kg bw/d in F0 parental animals.
Key result
Dose descriptor:
NOAEL
Remarks:
general, systemic toxicity
Effect level:
1 000 mg/kg bw/day (actual dose received)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
other: see 'Remark'
Key result
Critical effects observed:
no
Clinical signs:
no effects observed
Mortality / viability:
no mortality observed
Body weight and weight changes:
no effects observed
Sexual maturation:
not examined
Organ weight findings including organ / body weight ratios:
not examined
Gross pathological findings:
no effects observed
Histopathological findings:
no effects observed
The number of delivered F1 pups/litter, their postnatal survival and their body weights remained unaffected by administration of the test substance. Clinical and/or gross necropsy examinations of the F1 pups revealed only findings, which were considered to be spontaneous in nature.
Key result
Dose descriptor:
NOAEL
Remarks:
developmental toxicity
Generation:
F1
Effect level:
1 000 mg/kg bw/day (actual dose received)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
other: No adverse effects at 1000 mg/kg bw/d in F1 pups.
Key result
Critical effects observed:
no
Key result
Reproductive effects observed:
no

Parental animals

Blood chemistry (mean+/-SD (number of animals)) - statistical method: Kruskal-Wallis + Wilcoxon-test, Two sides, *p<0.05, ** p<0.010

 

 Male         

Female          

   0 mg/kg bw  100 mg/kg bw  300 mg/kg bw  1000 mg/kg bw  0 mg/kg bw  100 mg/kg bw  300 mg/kg bw  1000 mg/kg bw
 ALT µkat/l  0.49 +/-0.10 (5)  0.52 +/-0.09 (5)  0.49 +/-0.06 (5) 0.51 +/-0.11 (5)   0.46 +/-0.24 (5)  0.31 +/-0.07 (5)  0.30 +/-0.05 (5)  0.38 +/-0.09 (5)
 AST µkat/l  1.31 +/-0.30 (5)  1.46 +/-0.22 (5)  1.42 +/-0.23 (5)  1.32 +/-0.11 (5)  1.52 +/-0.18 (5)  1.38 +/-0.31 (5)  1.19 +/-0.20 (5)  1.20 +/-0.19 (5)
 ALP µkat/l  1.52 +/-0.27 (5)  1.46 +/-0.17 (5)  1.32 +/-0.17 (5)  1.53 +/-0.10 (5)  0.57 +/-0.16 (5)  1.66 +/-2.14 (5)  0.58 +/-0.07 (5)  0.71 +/-0.20 (5)
 SGGT nkat/l  0 +/-0 (5)

 0 +/-0 (5)

 0 +/-0 (5)  0 +/-0 (5)  0 +/-0 (5)  19 +/-42 (5)  0 +/-0 (5)  0 +/-0 (5)
 INP mmol/l  1.83 +/-0.06 (5)  1.91 +/-0.18 (5)  1.88 +/-0.06 (5)  2.06 +/-0.08** (5)  1.55 +/-0.15 (5)  1.64 +/-0.12 (5)  1.48 +/-0.17 (5)  1.74 +/-0.15 (5)
 Bilirubine µmol/l  1.96 +/-0.45 (5)  2.04 +/-0.23 (5)  2.13 +/-0.47 (5)  2.70 +/-0.62 (5)  2.51 +/-0.29 (5)  2.88 +/-0.28 (5)  2.76 +/-0.39 (5)  3.18 +/-0.43* (5)
 Protein 64.80 +/-1.86 (5)   63.86 +/-1.61 (5)  62.99 +/-2.20 (5)  62.54 +/-0.78 (5)  69.39 +/-1.43 (5)  67.35 +/-4.27 (5)  68.07 +/-1.71 (5)  64.12 +/-1.53** (5)
 Albumin  37.65 +/-0.98 (5)  36.59 +/-0.58 (5)  36.82 +/-1.41 (5)  36.57 +/-0.61 (5)  40.96 +/-0.87 (5)  38.99 +/-3.73 (5)  40.10 +/-0.77 (5)  37.50 +/-0.85** (5)
 Globulin  27.15 +/-1.04 (5)  27.27 +/-1.07 (5)  26.17 +/-1.48 (5)  25.97 +/-0.76 (5) 28.43 +/-1.10 (5)   28.53 +/-1.06 (5)  27.97 +/-1.07 (5)  26.62 +/-1.30 (5)
 Triglycerides  0.66 +/-0.45 (5)  0.52 +/-0.16 (5)  0.58 +/-0.34 (5)  0.55 +/-0.18 (5)  0.32 +/-0.05 (5)  0.32 +/-0.04 (5)  0.43 +/-0.10 (5)  0.42 +/-0.06* (5)
 Cholesterol  1.40 +/-0.19 (5)  1.74 +/-0.36* (5)  1.72 +/-0.14* (5)  1.47 +/-0.18 (5)  1.49 +/-0.22 (5)  1.55 +/-0.18 (5)  1.61 +/-0.32 (5)  1.57 +/-0.33 (5)
Conclusions:
The NOAEL for reproductive performance and fertility is 1000 mg/kg body weight/day for the F0 parental rats. The NOAEL for general, systemic toxicity of the test substance is 1000 mg/kg body weight/day for the F0 generation. The NOAEL for developmental toxicity is 1000 mg/kg bw/d. The NOAEL for developmental toxicity in the F1 progeny of the test substance treated groups was found to be 1000 mg/kg body weight/day.
Effect on fertility: via oral route
Endpoint conclusion:
no adverse effect observed
Dose descriptor:
NOAEL
1 000 mg/kg bw/day
Study duration:
subchronic
Species:
rat
Effect on fertility: via inhalation route
Endpoint conclusion:
no study available
Effect on fertility: via dermal route
Endpoint conclusion:
no study available
Additional information

BASF (2007) investigated the oral toxicity of the test substance after repeated exposure (54 days) in male/female Wistar rats. 12 males and 12 females (F0 animals) received daily (except animals being in labor) doses of 100, 300 and 1000 mg/kg bw/day. Most of the F0 parental animals proved to be fertile. The occurrence of one infertile pair each in test groups 1 and 3 (100 and 1000 mg/kg) did not suggest any relation to treatment. Gross and histopathological examinations of the respective animals of both genders did not reveal test substance-induced findings, which may have accounted for the observed infertility. Mating behavior, conception, gestation, parturition, lactation and weaning; sexual organ weights as well as gross and histopathological findings of these organs were comparable between the rats of the test substance-treated test groups and the corresponding controls and ranged within the historical control data of the test facility. Under the conditions of this reproductive/developmental toxicity screening test the NOAEL for reproductive performance and fertility is 1000 mg/kg body weight/day for the F0 parental rats.

Effects on developmental toxicity

Description of key information

Allt (2019) performed a pre-natal development toxicity study via oral gavage in female Sprague-Dawley rats according to OECD 414 (K1, GLP compliant). A NOAEL of 150 mg/kg bw/day was determined for the pregnant females, based on the absence of effects linked to systemic toxicity. No treatment related changes were detected in the offspring parameters measured or on embryofoetal development. The NOAEL for development toxicity was therefore considered to be 150 mg/kg bw/day.

Link to relevant study records
Reference
Endpoint:
developmental toxicity
Type of information:
experimental study
Adequacy of study:
key study
Study period:
04 April 2018 - 04 June 2018
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Qualifier:
according to
Guideline:
OECD Guideline 414 (Prenatal Developmental Toxicity Study)
Version / remarks:
adopted 22 January 2001
Deviations:
no
Qualifier:
according to
Guideline:
EPA OPPTS 870.3700 (Prenatal Developmental Toxicity Study)
Version / remarks:
August 1998
Deviations:
no
Qualifier:
according to
Guideline:
other: Japanese Ministry of Agriculture, Forestry and Fisheries Testing guidelines for Toxicology studies, 12 NohSan No 8147
Version / remarks:
24 November 2000
Deviations:
no
Qualifier:
according to
Guideline:
other: Commission Regulation (EC) No 440/2008 of 30 May 2008 test methods pursuant to Regulations (EC) No 1907/2006 of the European Parliament and of the Council on the Registration, Evaluation, Authorisation and Restriction of Chemicals (REACH)
Deviations:
no
GLP compliance:
yes (incl. certificate)
Limit test:
no
Specific details on test material used for the study:
SOURCE OF TEST MATERIAL
- Source and lot/batch No.of test material: TE767F05G
- Expiration date of the lot/batch: 05 June 2019
- Purity: 99.7%

STABILITY AND STORAGE CONDITIONS OF TEST MATERIAL
- Storage condition of test material: Room temperature in the dark
- Solubility and stability of the test substance in the solvent/vehicle: stable for 4 hours
- Samples were taken of test item formulations and were analyzed on two occasions for
concentration of Methoxypropylamine (MOPA) at Envigo Analytical Laboratory, Shardlow. The results indicate that the prepared formulations were within 7% of the nominal concentration.

OTHER SPECIFICS: No correction for purity was made.
Species:
rat
Strain:
Sprague-Dawley
Remarks:
Crl:CD (SD) IGS BR
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Charles River (UK) Limited, Margate, Kent
- Age at study initiation: no data
- Weight at study initiation: 199 to 285g
- Fasting period before study: no data
- Housing: The animals were housed in a single air-conditioned room within the Envigo Research Limited, Shardlow, UK Barrier Maintained Rodent Facility.
- Diet: ad libitum; A pelleted diet (Rodent 2018C Teklad Global Certified Diet, Envigo RMS (UK) Limited, Oxon, UK)
- Water: ad libitum; Mains drinking water was supplied from polycarbonate bottles attached to the cage.
- Acclimation period: no data

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22 ± 3 ºC
- Humidity (%): 50 ± 20%
- Air changes (per hr): at least fifteen air changes per hour
- Photoperiod (hrs dark / hrs light): twelve hours continuous light and twelve hours darkness.

IN-LIFE DATES: From: 07 April 2018 (first day of treatment) and 26 April 2018 (final day of necropsy).
Route of administration:
oral: gavage
Vehicle:
arachis oil
Details on exposure:
PREPARATION OF DOSING SOLUTIONS: Formulations were shown to be stable for four hours. Formulations were therefore prepared daily and dosed within four hours of preparation.

VEHICLE
- Concentration in vehicle: 7.5, 18.75 and 37.5 mg/mL
- Treatment volume: 4 mL/kg
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
The test item concentration in the test samples was determined by gas chromatography (GC) using an external standard technique. The test item gave a chromatographic profile consisting of a single peak.
The analytical procedure was successfully validated with respect to specificity of chromatographic analysis, linearity of detector response, method accuracy and precision.
The homogeneity and stability was confirmed for test item in Arachis Oil BP formulations at nominal concentrations of 2.5 mg/mL and 100 mg/mL when stored for 4 hours during study number WD30CK.
The mean concentrations of test item in test formulations analyzed for the study were within ± 10% of nominal concentrations, confirming accurate formulation.
Details on mating procedure:
Animals were delivered in two batches containing females prior to Day 3 of gestation. The day that positive evidence of mating was observed was designated Day 0 of gestation.
Duration of treatment / exposure:
between Days 3 and 19 of gestation
Frequency of treatment:
daily
Duration of test:
from Day 3 to 19 of gestation
Dose / conc.:
0 mg/kg bw/day (nominal)
Remarks:
Control group
Dose / conc.:
30 mg/kg bw/day (nominal)
Remarks:
low treatment group
Dose / conc.:
75 mg/kg bw/day (nominal)
Remarks:
Intermediate treatment group
Dose / conc.:
150 mg/kg bw/day (nominal)
Remarks:
high treatment group
No. of animals per sex per dose:
24 females per group
Control animals:
yes, concurrent vehicle
Details on study design:
- Dose selection rationale: The dose levels were chosen in collaboration with the Sponsor Representative and were based on available toxicity data including a Preliminary Oral (Gavage) Pre-Natal Development Toxicity Study in the Rat (Covance CRS Study Number VF78GT) and a 90 Day Oral (Gavage) Toxicity Study in the Rat (Covance CRS Study Number WD30CK).
- Rationale for animal assignment: The animals were randomly allocated to treatment groups using a randomization procedure based on stratified body weight to ensure similarity between the treatment groups.
Maternal examinations:
DETAILED CLINICAL OBSERVATIONS: Yes
Following arrival, all animals were examined for overt signs of toxicity, ill-health or behavioral changes once daily during the gestation period. Additionally, during the dosing period, observations were recorded immediately before and soon after dosing and one hour post dosing. All observations were recorded.

BODY WEIGHT: Yes
Individual body weights were recorded on Day 3 (before the start of treatment) and on Days 4, 5, 8, 11, 14 and 17 of gestation. Body weights were also recorded for animals at terminal kill (Day 20).

FOOD CONSUMPTION: Yes
Food consumption was recorded for each individual animal at Day 3, 5, 8, 11, 14, 17 and 20 of gestation.

WATER CONSUMPTION: Yes
Water intake was observed daily by visual inspection of the water bottles for any overt changes.

POST-MORTEM EXAMINATIONS: Yes
All animals were killed by carbon dioxide asphyxiation followed by cervical dislocation on Day 20 of gestation. All animals were subjected to a full external and internal examination and any macroscopic abnormalities were recorded.
Ovaries and uterine content:
The ovaries and uteri of pregnant females were removed, examined and the following data recorded:
ii) Number, position and type of intrauterine implantation
iii) Fetal sex
iv) External fetal appearance
v) Fetal weight
vi) Placental weight
vii) Gravid uterus weight
The uteri of any apparently non-pregnant females were immersed in 0.5% ammonium polysulphide solution to reveal evidence of implantation.

Implantation types were divided into:
Early Death: No visible distinction between placental/decidual tissue and embryonic tissue
Late Death: Separate embryonic/fetal and placental tissue visible
Dead Fetus: A fetus that had died shortly before necropsy. These were included as late deaths for reporting purposes.

All implantations and viable fetuses were numbered according to their intrauterine position as follows (as an example):
Left Horn Cervix Right Horn
L1 L2 L3 L4 L5 L6 L7 L8 R1 R2 R3 R4 R5 R6 R7 R8
V1 V2 V3 V4 V5 V6 V7 V8 V9 V10 V11 V12 V13 V14 V15 V16
V = viable fetus
Fetal examinations:
The fetuses were killed by subcutaneous injection of a suitable barbiturate agent. Fetuses from each litter were divided into two groups and examined for skeletal alterations and soft tissue alterations. Alternate fetuses were identified using an indelible marker and placed in Bouin’s fixative. Fetuses were subsequently transferred to distilled water and examined for visceral anomalies under a low power binocular microscope and then stored in 10% Buffered Formalin. The remaining fetuses were identified using cardboard tags marked with chinagraph pencil and placed into 70% IMS in distilled water.
The fetuses were subsequently eviscerated, processed and the skeletons stained with alizarin red S before being transferred to 50% glycerol for examination of skeletal development and anomalies and storage.
Statistics:
Data was assessed using the R Environment for Statistical Computing. Initially, the distribution of the data was assessed by the Shapiro-Wilk normality test, followed by assessment of the homogeneity of the data using Bartlett’s test. Where considered appropriate, parametric analysis of the data was applied incorporating analysis of variance (ANOVA), which if significant, was followed by pair-wise comparisons using Dunnett’s test. Where parametric analysis of the data was considered to be unsuitable, non-parametric analysis of the data was performed incorporating the Kruskal-Wallis test which if significant was followed by the Mann-Whitney "U" test. Dose response relationships were also investigated by linear regression.
Probability values (p) are presented as follows:
p<0.001 ***
p<0.01 **
p<0.05 *
p≥0.05 (not significant)
Indices:
Percentage pre-implantation loss was calculated as:
[(number of corpora lutea-number of implantations)/number of corpora lutea] x 100

Percentage post-implantation loss was calculated as:
[(number of implantations - number of live fetuses)/number of implantations] x 100

Sex ratio was calculated as:
% male fetuses (sex ratio) = (number of male fetuses/total number of fetuses) x 100
Clinical signs:
effects observed, non-treatment-related
Description (incidence and severity):
Instances of increased salivation were evident in all females treated with 150 mg/kg bw/day between Days 13 and 19 and in two females treated with 75 mg/kg bw/day on Day 19 only.
No such effects were detected in females treated with 30 mg/kg bw/day.

The following observations were noted in a small number of control or treated females and in isolation, these were considered to be incidental and unrelated to the test item. One control female had scattered scabs on one occasion, another control female had red/brown staining around the eyes on one occasion, one female treated with 75 mg/kg bw/day had red/brown staining around the mouth on one occasion, and three females treated with 150 mg/kg bw/day had generalized fur loss between Days 16 and 20, with one of these females also having red/brown staining around the mouth on one occasion.
Dermal irritation (if dermal study):
not examined
Mortality:
no mortality observed
Description (incidence):
There were no unscheduled deaths.
Body weight and weight changes:
no effects observed
Description (incidence and severity):
Body weight and body weight gain, including adjustment for the contribution of the gravid uterus, was unaffected by treatment at 30, 75 or 150 mg/kg bw/day.

Females from all treatment groups showed a statistically significant increase (p<0.05-0.001) in body weight gain between Days 3 and 4 of gestation. An increase in body weight gain is considered not to reflect an adverse effect of treatment and the increase was not dose related and was therefore considered not to be of toxicological significance.
Food consumption and compound intake (if feeding study):
no effects observed
Description (incidence and severity):
No adverse effect on food consumption was evident in females from any treatment group.

Females treated with 150 mg/kg bw/day exhibited a statistically significant reduction (p<0.01) in food consumption between Days 8 and 11 of gestation. As no such effects were evident before or after this period and all but two individual values for food consumption at this treatment group were within the historical control data range, this reduction was considered not to be of any toxicological significance.
Food efficiency:
not examined
Water consumption and compound intake (if drinking water study):
no effects observed
Description (incidence and severity):
Daily visual inspection of water bottles did not reveal any overt intergroup differences.
Ophthalmological findings:
not examined
Haematological findings:
not examined
Clinical biochemistry findings:
not examined
Urinalysis findings:
not examined
Behaviour (functional findings):
not examined
Immunological findings:
not examined
Organ weight findings including organ / body weight ratios:
not examined
Gross pathological findings:
effects observed, treatment-related
Description (incidence and severity):
Sixteen females treated with 150 mg/kg bw/day had ulceration on the non-glandular region of the stomach. One of these females also had a pale stomach. A further five females from this treatment group had ulceration on both the non-glandular and glandular regions of the stomach. Eleven females treated with 75 mg/kg bw/day had ulceration on the non-glandular region of the stomach and one female treated with 30 mg/kg bw/day had white ridges on the non-glandular region of the stomach.
One female treated with 150 mg/kg bw/day had a mass on the right side of the abdominal cavity. In isolation this was considered to be incidental and unrelated to treatment.
Neuropathological findings:
not examined
Histopathological findings: non-neoplastic:
not examined
Histopathological findings: neoplastic:
not examined
Number of abortions:
no effects observed
Pre- and post-implantation loss:
no effects observed
Description (incidence and severity):
There was no treatment-related effect observed in the pre- and post-implantation loss in all dose groups, when comparing to the control group:
- preimplantation loss: 15.6%, 18.1%, 17.1%, 18.7% at 0, 30, 75 and 150 mg/kg bw/day
- post-implantation loss: 2.3%, 1.3%, 1.5%, 2.7% at 0, 30, 75 and 150 mg/kg bw/day
All data was within the historical control range available for this species and strain.
Total litter losses by resorption:
no effects observed
Description (incidence and severity):
There were no total litter losses by resorption observed.
Early or late resorptions:
no effects observed
Description (incidence and severity):
There were no treatment-related effects on early or late resorptions observed.
Dead fetuses:
no effects observed
Description (incidence and severity):
There were no treatment-related effects on the number of dead fetuses observed.
Changes in pregnancy duration:
no effects observed
Description (incidence and severity):
Cumulative Body Weight Changes were recorded until day 20 of pregnancy, suggesting a duration of 20 days for all females.
Changes in number of pregnant:
no effects observed
Description (incidence and severity):
There was no treatment-related, toxicologically relevant effect on the number of pregnant dams per dose group. The number of pregnant females per dose group were 24/24, 24/24, 24/24 and 24/24 at 0,25, 100 and 250 mg/kg bw/day, resp. No historical control data is available for this parameter.
Details on maternal toxic effects:
There was no obvious effect of maternal treatment on litter data as assessed by numbers of implantations, in-utero offspring survival (as assessed by the mean numbers of early or late resorptions), live litter size, sex ratio and pre and post-implantation losses at 30, 75 or 150 mg/kg bw/day.
Intergroup differences for mean fetal, litter or placental weights did not indicate any adverse effects of maternal treatment at 30, 75 or 150 mg/kg bw/day.
Key result
Dose descriptor:
NOAEL
Effect level:
150 mg/kg bw/day (nominal)
Based on:
test mat.
Basis for effect level:
clinical signs
gross pathology
Remarks on result:
other: at a lesser extent, also evident in females treated with 75 mg/kg bw/day
Key result
Abnormalities:
no effects observed
Fetal body weight changes:
effects observed, non-treatment-related
Description (incidence and severity):
Intergroup differences for mean fetal, litter or placental weights did not indicate any adverse effects of maternal treatment at 30, 75 or 150 mg/kg bw/day.
Male fetal weight and combined fetal weights for litters from females treated with 30 mg/kg bw/day showed a statistically significant increase (p<0.05-0.01) when compared to controls. An increase in fetal weight is generally not considered to represent an adverse effect of treatment and in the absence of a similar effect at the higher dosages, the intergroup differences are considered not to be of toxicological significance.
Reduction in number of live offspring:
no effects observed
Description (incidence and severity):
There is no treatment-related effect on the number of live offspring.
Changes in sex ratio:
no effects observed
Description (incidence and severity):
There was no treatment-related effect observed on sex ratio.
Changes in litter size and weights:
no effects observed
Description (incidence and severity):
There was no treatment-related effects observed on litter weight or litter size.
Changes in postnatal survival:
no effects observed
Description (incidence and severity):
There was no treatment-related effects observed on postnatal survival.
External malformations:
no effects observed
Description (incidence and severity):
Neither the type, incidence nor distribution of external finding apparent for fetuses at Day 20 of gestation indicated an adverse effect of maternal treatment on fetal development at 30, 75 or 150 mg/kg bw/day.

A statistically significant increase (p<0.05) in the number of fetuses/litters showing a large placenta was evident at 150 mg/kg bw/day. The group mean value (4.7%) was slightly above the historical control range (0.0 - 4.2%), however, this was considered to be the result of one litter which had five fetuses that had this finding. With this litter excluded, the group mean value was within the historical control range. In the absence of any associated effects, the intergroup difference was considered not to be of toxicological significance.
Skeletal malformations:
effects observed, non-treatment-related
Description (incidence and severity):
Skeletal examinations of fetuses on Day 20 of gestation did not indicate any obvious effect of maternal treatment on fetal development at 30, 75 or 150 mg/kg bw/day.
Statistically significant increases (p<0.01) in the number of fetuses/litters showing a ossification centre associated with the first lumbar vertebra, incomplete ossification of the squamosal of the skull and zygomatic process of maxilla of the skull were noted in females treated with 150 mg/kg bw/day. The observations of one variant at a higher incidence compared with controls is not significant when evaluated in isolation. In the absence of a particular pattern of abnormal skeletal development of skeletal structures affecting treated fetuses, the observation of one affected skeletal structure can be considered unlikely to represent true developmental abnormality. This also takes account of these findings being seen regularly on this study type amongst control group fetuses.
A statistically significant increase (p<0.05) in the number of fetuses/litter showing incomplete ossification of the sternebra was evident at 75 mg/kg bw/day. Statistically significant reductions (p<0.05-0.01) in incomplete ossification of the cervical (neural) arch and incomplete ossification of the occipital (supra-occipital) was evident at 30 mg/kg bw/day. In the absence of similar findings at higher dosages or any particular pattern of abnormal skeletal development of skeletal structures affecting treated fetuses, the observation of one affected skeletal structure can be considered unlikely to represent true developmental abnormality.
Visceral malformations:
effects observed, non-treatment-related
Description (incidence and severity):
Visceral examinations of fetuses on Day 20 of gestation did not indicate any obvious effect of maternal treatment on fetal development at 30, 75 or 150 mg/kg bw/day.

A statistically significant increase (p<0.05) in the number of fetuses/litter showing a nonuniform patterning of the rugae was evident at 75 and 30 mg/kg bw/day. In the absence of a similar finding at the highest dosage, the intergroup differences were considered unrelated to treatment.
Details on embryotoxic / teratogenic effects:
No treatment-related changes were detected in the offspring parameters measured or on embryofetal development. The ‘No Observed Effect Level’ (NOEL) for developmental toxicity was therefore considered to be 150 mg/kg bw/day.
Key result
Dose descriptor:
NOAEL
Effect level:
150 mg/kg bw/day (nominal)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
other: developmental toxicity
Key result
Abnormalities:
no effects observed
Key result
Developmental effects observed:
no

Table 1   Summary of Female Performance

Category

Number of Females at Dose Level (mg/kg bw/day)

0 (Control)

30

75

150

Initial Group Size

24

24

24

24

Pregnant

24

24

24

24

 

Table 2   Summary Incidence of Daily Clinical Observations

Dose Level

(mg/kg bw/day)

Number of Animals

Clinical Observations

Number Showing Effect (Days post coitum affected)

0 (Control)

24

No Abnormalities Detected

Scattered Scabs

22 (0-20)

1 (5)

 

 

Red/Brown Staining around the Eyes

1 (9)

30

24

No Abnormalities Detected

24 (0-20)

75

24

No Abnormalities Detected

Increased Salivation

21 (0-20)

2 (19)

 

 

Red/Brown Staining Around the Mouth

1 (19)

150

24

No Abnormalities Detected

Fur Loss (Hind Quarters)

3 (0-20)

3 (16-20)

 

 

Red/Brown Staining Around the Mouth

1 (13)

 

 

Increased Salivation

20 (13-19)

 

Table 3   Group Mean Body Weight Values

Dose Level

(mg/kg bw/day)

 

3

4

Body Weight (g) on Day of Gestation

5          8          11       14

17

20

0 (Control)

mean

sd

260.7

21.6

261.6

20.9

267.0

20.2

280.6

22.9

300.8

23.1

320.0

25.0

347.4

27.4

395.8

33.6

 

n

24

24

24

24

24

24

24

24

30

mean

sd

258.6

16.2

261.3

17.4

266.4

17.9

280.0

19.5

300.1

21.0

318.7

23.2

349.5

23.8

393.9

30.6

 

n

24

24

24

24

24

24

24

24

75

mean

sd

256.8

15.8

261.7

15.5

264.7

16.2

276.6

16.5

295.9

17.3

314.8

18.6

342.8

22.1

388.9

28.7

 

n

24

24

24

24

24

24

24

24

150

mean

sd

255.8

16.3

260.5

16.4

264.2

16.9

276.1

15.6

293.9

17.3

312.6

17.7

339.6

21.5

384.5

26.7

 

n

24

24

24

24

24

24

24

24

 

Table 4   Group Mean Body Weight Change Values 

Dose

Level

 

(mg/kg bw/day)

3 to 4

 

Body Weight Change (g) during Days of Gestation

4 to 5   5 to 8   8 to 11    11 to 14   14 to 17

17 to 20

0

(Control)

mean

sd

0.9

5.6

 

5.4

3.7

13.6

5.3

20.2

4.8

19.2

6.1

27.4

5.3

48.4

8.1

 

n

24

 

24

24

24

24

24

24

30

mean

sd

2.8*

3.8

 

5.1 3.9

13.6

4.6

20.1

4.4

18.6

6.5

30.8

5.3

44.4

9.2

 

n

24

 

24

24

24

24

24

24

75

mean

sd

4.9***

2.9

 

3.0 3.2

11.9

4.4

19.3

4.7

18.9

5.5

28.0

6.6

46.1

8.3

 

n

24

 

24

24

24

24

24

24

150

mean

sd

4.7*

7.8

 

3.7 3.6

11.9

4.1

17.8

5.5

18.7

5.2

27.0

8.2

44.9

7.7

 

n

24

 

24

24

24

24

24

24

 

Dose Level

(mg/kg bw/day)

 

4

Cumulative Body Weight Change (g) from Day 3 of Gestation

           5                8               11              14              17

20

0 (Control)

mean

sd

0.9 5.6

6.3

5.1

19.9

7.9

40.1

8.6

59.3

10.7

86.7

13.2

135.1

19.3

 

n

24

24

24

24

24

24

24

30

mean

sd

2.8*

3.8

7.8

4.5

21.4

6.7

41.5

8.4

60.1

11.5

90.9

12.3

135.3

19.8

 

n

24

24

24

24

24

24

24

75

mean

sd

4.9***

2.9

7.9

3.2

19.8

5.6

39.1

6.2

58.0

7.9

86.0 10.1

132.1

16.6

 

n

24

24

24

24

24

24

24

150

mean

sd

4.7*

7.8

8.4

8.1

20.3

9.1

38.1

10.9

56.8

12.2

83.8

15.8

128.7

21.4

 

n

24

24

24

24

24

24

24

Table 5        Group Mean Gravid Uterus Weight and Adjusted Body Weight and Body Weight Change Values

Dose Level

(mg/kg bw/day)

 

Body Weight (g) on Days of Gestation

 3            20

Body Weight

Change (g) during Days of Gestation

3-20

Gravid

Uterus

Weight

(g)

Adjusted

Body

Weight (g) Day 20

Adjusted

Body Weight

Change (g)

3-20

0 (Control)

mean

sd

260.7

21.6

395.8

33.6

135.1

19.3

80.985 13.696

314.8

27.1

54.1 11.5

 

n

24

24

24

24

24

24

30

mean

sd

258.6

16.2

393.8

30.4

135.2

19.5

78.509 14.164

315.3

25.8

56.7 15.9

 

n

24

24

24

24

24

24

75

mean

sd

256.8

15.8

388.9

28.7

132.1

16.6

80.375 12.871

308.5

23.0

51.8 11.4

 

n

24

24

24

24

24

24

150

mean

sd

255.8

16.3

384.5

26.7

128.7

21.4

77.238 19.951

307.3

19.3

51.5 12.2

 

n

24

24

24

24

24

24

 

Table 6   Group Mean Food Consumption Values

Dose Level

(mg/kg bw/day)

 

Food Consumption (g/rat/day) between Days of Gestation

3 - 5        5 - 8       8 - 11     11 - 14    14-17      17 - 20

0 (Control)

mean

sd

21.4

1.8

23.0

2.5

24.2

2.2

24.3

2.4

25.3

4.8

27.3

4.5

 

n

24

23

24

24

24

24

30

mean

sd

21.0

2.8

23.2

3.2

23.7

3.0

24.4

6.0

26.5

5.1

26.3

3.4

 

n

24

24

24

24

24

24

75

mean

sd

21.5

3.3

24.2

10.0

24.1

6.4

23.2

2.7

26.0

4.9

26.2

3.6

 

n

24

24+

23#

24

24

24

150

mean

sd

20.6

4.6

21.2

2.0

21.7**

2.5

22.6

1.9

24.1

4.9

26.0

5.1

 

n

24

24

24

24

24

24

   + = Food consumption for Female 61 calculated for Days 5-7 only due to female being present in incorrect cage on Day 8. 

# = Data unavailable for Female No. 63, due to erroneous value recorded for food allocated on Day 8. 

Table 7   Summary Incidence of Necropsy Findings

 

 

Dose Level (mg/kg bw/day)

 

0 (Control)

30

75

150

TERMINAL DEATH

 

Number of Animals Examined

 

 

24

 

 

24

 

 

24

 

 

24

 

External:

Generalized Fur Loss

 

 

0

 

 

0

 

 

0

 

 

1

Mass on Right Side

0

0

0

1

 

Internal:

Stomach – Non-Glandular Region: White Ridges

 

 

0

 

 

1

 

 

0

 

 

0

Stomach – Non-Glandular Region: Ulcerated

0

0

11

16

Stomach: Ulcerated

0

0

0

5

Stomach: Pale

0

0

0

1

Outer Right Side of Abdominal Cavity: Mass

0

0

0

1

 

No Abnormalities Detected

 

 

24

 

 

23 

 

13

 

 

3

 

 


Table 8   Group Mean Litter Data Values

Dose Level

(mg/kg bw/day)

 

Number of

Corpora Lutea

Number of

Implants

Number of

Embryonic/Fetal Deaths

Implantation

Loss 

%

Number of Live Implants

%

Male

Fetuses

Mean

Male

Fetal

Weight

(g)

Mean

Female Fetal

Weight

(g)

Mean

Fetal

Weight

(g)

Mean

Placental

Weight

(g)

Litter

Weight

(g)

Total

Placental

Weight

(g)

Early

Late

Total

Pre

Post

Male

Female

Total

0 (Control)

mean

sd

16.3

2.3

13.8

2.6

0.2

0.5

0.1

0.3

0.3 0.6

15.6 12.3

2.3 3.8

6.7

2.2

6.7

2.0

13.4

2.5

50.3

13.0

3.917

0.315

3.742

0.306

3.831

0.286

0.537

0.065

51.054

9.067

7.137

1.459

 

n

23#

24

24

24

24

23

24

24

24

24

24

24

24

24

23+

24

23+

30

mean

sd

15.4

2.4

12.8

2.8

0.1 0.3

0.0 0.2

0.2 0.4

18.1 13.4

1.3 3.0

6.3 1.8

6.3 2.2

12.6

2.9

52.1

15.2

4.253**

0.546

3.906

0.271

4.140*

0.569

0.571 0.132

50.878

9.780

6.834

1.202

 

n

24

24

24

24

24

24

24

24

24

24

24

24

23

24

24+

24

24+

75

mean

sd

16.2

2.3

13.5

2.5

0.2

0.4

0.0 0.2

0.2 0.4

17.1

7.4

1.5 3.0

7.1 2.7

6.2 2.1

13.3

2.5

53.0

17.2

3.961

0.291

3.771

0.305

3.877 0.279

0.540 0.045

51.140

8.766

7.138

1.334

 

n

24

24

24

24

24

24

24

24

24

24

24

24

24

24

24

24

24

150

mean

sd

15.3

2.2

12.6

3.2

0.1

0.4

0.0

0.2

0.2

0.5

18.7

14.7

2.7

10.3

6.5

2.5

5.9

2.5

12.4

3.4

52.9

14.3

4.029

0.264

3.781

0.309

3.913

0.289

0.608

0.165

48.735

13.796

7.201

1.968

 

n

24

24

24

24

24

24

24

24

24

24

24

24

24

24

24

24

24

+ = Placental weights not recorded in error for Female No. 21’s litter and for fetus No. 8 from Female No. 25

# = Data unavailable for Female No. 16

Table 9   Summary Incidence of Fetal External Findings

 

External Findings

 

Dose level (mg/kg bw/day)

 

 

0 (Control)

30

75

 

150

 

Number of fetuses (litters) examined

 

 

322 (24)

303 (24)

318 (24)

 

298 (24)

NF

NL

%†

NF

NL

%†

NF

NL

%†

NF

NL

%†

Total Number Affected

 

4

1

1.3

8

5

5.9

7

6

2.2

15

6

6.6

Small Fetus

 

4

1

1.3

6

4

1.8

5

4

1.6

8

4

2.8

Pale Fetus

 

1

1

0.3

0

0

0.0

0

0

0.0

0

0

0.0

Large Fetus

 

0

0

0.0

2

1

4.2

0

0

0.0

0

0

0.0

Large Placenta

 

0

0

0.0

2

1

4.2

1

1

0.3

9

5

4.7*

Small Placenta

 

2

1

0.6

1

1

0.2

2

2

0.7

1

1

0.3

 

            

Table 10 Summary Incidence of Fetal Visceral Findings

Visceral Findings

 

 

 

 

Dose Level (mg/kg bw/day)

 

 

 

 

 

0 (Control)

 

30

75

 

 

150

 

 

 

 

 

Number of Fetuses (litters) Examined

 

 

 

 

 

168 (24)

 

 

156 (24)

164 (24)

 

 

157 (24)

 

NF

 

NL

%†

NF

NL

%†

NF

NL

%†

NF

NL

%†

External

Hemorrhage 

 

0

 

 

0

 

0.0

 

0

 

0

 

0.0

 

1

 

1

 

0.5

 

0

 

0

 

0.0

Head

Rugae - non-uniform patterning

 

2

 

 

2

 

1.0

 

11

 

8

 

6.8*

 

8

 

8

 

4.8*

 

5

 

2

 

3.9

Abdomen

Liver - additional lobe between right and left median

 

2

 

 

2

 

1.2

 

2

 

1

 

1.0

 

0

 

0

 

0.0

 

0

 

0

 

0.0

Umbilical artery - left-sided

0

 

0

0.0

0

0

0.0

1

1

0.8

2

2

1.2

Testis - partially undescended

4

 

3

2.4

4

4

2.5

1

1

0.6

2

2

1.2

Testis - malrotated

0

 

0

0.0

0

0

0.0

0

0

0.0

1

1

0.5

Ureter - kinked

15

 

10

10.0

11

7

7.0

12

6

8.3

9

7

7.3

Ureter - dilated 

11

 

7

7.9

3

3

1.9

8

4

5.9

5

4

4.9

Kidney - malpositioned 

0

 

0

0.0

0

0

0.0

1

1

0.8

0

0

0.0

Renal pelvic cavitation - increased 

8

 

6

4.4

8

7

5.0

7

5

4.5

10

6

7.5

Renal papilla - absent

1

 

1

0.5

2

2

1.3

3

2

2.3

1

1

1.4

Renal medulla - reduced in size

1

 

1

0.5

0

0

0.0

0

0

0.0

0

0

0.0

Thorax

Thymus - lobe partially undescended

 

5

 

 

4

 

2.8

 

5

 

4

 

2.9

 

3

 

3

 

1.8

 

4

 

4

 

2.2

Atrium - enlarged

4

 

4

2.3

1

1

0.7

2

2

1.1

3

3

1.6

Ventricle - reduced in size

0

 

0

0.0

0

0

0.0

1

1

0.5

0

0

0.0

Ventricle - vacuole

0

 

0

0.0

0

0

0.0

1

1

0.5

0

0

0.0

Total

30

 

18

18.5

31

14

18.9

27

14

17.0

30

15

20.3

 

Table 11 Summary Incidence of Fetal Skeletal Findings

Skeletal Findings

 

Dose Level (mg/kg bw/day)

 

 

0 (Control)

30

75

 

150

 

Number of Fetuses (litters) Examined

 

 

154 (24)

147 (24)

154 (24)

 

141 (24)

NF

NL

%†

NF

NL

%†

NF

NL

%†

NF

NL

%†

Skull

Fontanelle (anterior) - large

 

2

 

1

 

1.4

 

2

 

2

 

1.1

 

9

 

4

 

5.2

 

1

 

1

 

0.6

Nasal - incomplete ossification

10

5

6.2

6

5

3.9

18

9

10.5

19

9

12.5

Nasal/Frontal - sutural bone

1

1

0.7

0

0

0.0

0

0

0.0

0

0

0.0

Frontal - incomplete ossification

1

1

0.7

0

0

0.0

5

4

2.9

8

5

5.1

Frontal - unossified area

1

1

0.7

3

2

1.9

5

3

3.5

3

2

1.6

Parietal - incomplete ossification

6

4

3.7

3

2

2.1

8

6

5.0

20

10

14.2

Interparietal - incomplete ossification

29

11

18.4

14

8

9.1

30

12

18.5

38

14

25.7

Occipital (Supra-occipital) - incomplete ossification

17

12

11.1

6

4

4.1*

23

10

14.2

26

14

18.4

Occipital (Supra-occipital) - unossified area(s)

12

8

7.2

7

6

4.3

8

5

4.6

8

7

6.5

Squamosal - incomplete ossification

7

5

4.4

8

4

5.4

14

7

8.1

30

14

20.3**

Squamosal - unossified area(s)

1

1

0.8

0

0

0.0

1

1

0.5

1

1

1.4

Jugal - incomplete ossification

5

3

3.2

4

4

2.6

4

4

2.5

8

6

6.7

Zygomatic process of maxilla - incomplete ossification

4

3

2.6

7

6

4.3

13

9

7.6

18

11

12.5**

Zygomatic process of squamosal - incomplete ossification

1

1

0.7

0

0

0.0

2

2

1.1

6

3

4.2

Zygomatic process of squamosal - fused to jugal

1

1

0.7

0

0

0.0

0

0

0.0

0

0

0.0

Premaxilla - incomplete ossification

0

0

0.0

0

0

0.0

3

3

1.8

2

1

1.4

Hyoid - incomplete ossification

14

9

8.6

17

11

11.9

11

8

7.0

16

9

10.1

Hyoid - not ossified

14

10

9.1

5

5

3.6

11

6

6.4

14

7

8.3

Presphenoid - incomplete ossification

1

1

0.7

0

0

0.0

1

1

0.7

2

2

1.3

Presphenoid - not ossified

0

0

0.0

1

1

0.7

2

2

1.3

0

0

0.0

NOTE: a fetus may appear in more than one category


Skeletal Findings

Dose Level (mg/kg bw/day)

 

 

0 (Control)

30

75

 

150

 

Number of Fetuses (litters) Examined

 

 

154 (24)

147 (24)

154 (24)

 

141 (24)

 

NF

NL

%†

NF

NL

%†

NF

NL

%†

NF

NL

%†

Vertebral Column

Odontoid - ossification present

 

0

 

0

 

0.0

 

1

 

1

 

4.2

 

0

 

0

 

0.0

 

1

 

1

 

0.7

Ventral arch of vertebra 1 - ossification present

45

18

31.7

49

18

36.3

37

16

23.9

34

11

22.2

Cervical (neural) arch - incomplete ossification

8

7

5.1

0

0

0.0**

10

7

6.3

16

9

10.1

Thoracic centrum - incomplete ossification

12

9

8.1

3

2

1.8

10

5

6.3

7

6

5.0

Thoracic centrum - not ossified

3

1

2.1

0

0

0.0

1

1

0.8

2

2

1.3

Thoracic centrum - bipartite ossification

5

4

3.8

2

2

1.4

2

2

1.1

1

1

0.8

Thoracic centrum - dumb-bell-shaped

20

10

13.9

19

11

16.2

19

13

11.7

21

14

18.5

Thoracic centrum - asymmetrically ossified

4

4

3.0

1

1

0.6

2

2

1.1

3

3

2.2

Thoracic centrum - misaligned

1

1

0.8

0

0

0.0

0

0

0.0

0

0

0.0

Thoracic centrum - hemicentric 

2

2

1.5

0

0

0.0

0

0

0.0

0

0

0.0

Thoracic vertebra - absent (arch and centrum)

1

1

0.8

0

0

0.0

0

0

0.0

0

0

0.0

Lumbar centrum - incomplete ossification

3

2

2.1

0

0

0.0

0

0

0.0

0

0

0.0

Lumbar centrum - bipartite ossification

0

0

0.0

0

0

0.0

1

1

0.8

0

0

0.0

Lumbar centrum - dumb-bell-shaped

0

0

0.0

0

0

0.0

1

1

0.8

0

0

0.0

Lumbar centrum - asymmetrically ossified

0

0

0.0

0

0

0.0

1

1

0.8

0

0

0.0

Sacral centrum - incomplete ossification

0

0

0.0

0

0

0.0

1

1

0.8

0

0

0.0

Sacral centrum - bipartite ossification

0

0

0.0

0

0

0.0

1

1

0.8

0

0

0.0

Sacral (neural) arch - incomplete ossification

25

11

15.9

22

9

15.3

31

13

19.9

39

14

26.5

Sacral (neural) arch - not ossified

0

0

0.0

0

0

0.0

1

1

0.6

0

0

0.0

Caudal vertebrae - less than 4 ossified

28

14

18.0

24

12

14.8

47

13

28.0

51

15

32.8

Number of pre-sacral vertebrae = 25/27 

1

1

0.8

1

1

0.7

1

1

0.8

1

1

2.1

 

Skeletal Findings

Dose Level (mg/kg bw/day)

 

0 (Control)

30

75

 

150

Number of Fetuses (litters) Examined

 

154 (24)

147 (24)

154 (24)

 

141 (24)

NF

NL

%†

NF

NL

%†

NF

NL

%†

NF

NL

%†

Ribs

Ossification centre - associated with 7th cervical vertebra

 

0

 

0

 

0.0

 

1

 

1

 

0.8

 

0

 

0

 

0.0

 

2

 

2

 

1.4

14th rib - extra - associated with 1st lumbar vertebra

0

0

0.0

1

1

0.7

0

0

0.0

0

0

0.0

Ossification centre - associated with 1st lumbar vertebra

2

2

1.5

8

4

5.5

4

3

2.5

18

11

14.1**

One or more ribs - wavy

0

0

0.0

0

0

0.0

0

0

0.0

2

1

1.0

One or more ribs - thickened

1

1

0.6

0

0

0.0

0

0

0.0

2

1

1.0

Rib - short

3

1

1.8

1

1

0.7

2

2

1.5

0

0

0.0

Rib - fused 

2

2

1.5

0

0

0.0

0

0

0.0

0

0

0.0

Rib - incomplete ossification

0

0

0.0

0

0

0.0

0

0

0.0

1

1

0.5

Rib - interrupted ossification

1

1

0.8

0

0

0.0

0

0

0.0

0

0

0.0

Rib - not ossified

1

1

0.8

0

0

0.0

0

0

0.0

0

0

0.0

Rib - absent

1

1

0.8

0

0

0.0

0

0

0.0

0

0

0.0

Rib - no articulation point

1

1

0.8

0

0

0.0

1

1

0.5

0

0

0.0

Rib - bifurcated

1

1

0.8

0

0

0.0

0

0

0.0

0

0

0.0

Costal cartilage - misaligned

3

2

2.0

2

2

1.2

3

3

1.6

1

1

0.7

Costal cartilage - not fused to sternebra

14

9

8.7

11

9

7.5

12

7

7.9

14

7

11.1

Sternebrae

Sternebra - incomplete ossification

 

4

 

2

 

2.7

 

0

 

0

 

0.0

 

11

 

8

 

6.2*

 

4

 

3

 

2.7

Sternebra - not ossified

3

1

2.1

0

0

0.0

1

1

0.7

0

0

0.0

Sternebra - bipartite ossification

0

0

0.0

0

0

0.0

0

0

0.0

1

1

0.7

Sternebra - misaligned

2

2

1.3

4

4

2.4

6

5

3.3

8

6

4.9

Sternum - split

0

0

0.0

0

0

0.0

1

1

0.7

0

0

0.0

 

Skeletal Findings

Dose Level (mg/kg bw/day)

 

 

0 (Control)

30

75

 

150

 

Number of Fetuses (litters) Examined

 

 

154 (24)

147 (24)

154 (24)

 

141 (24)

 

 

 

NF

NL

%†

NF

NL

%†

NF

NL

%†

NF

NL

%†

Sternebrae (continued)

Xiphoid cartilage - split

 

0

 

0

 

0.0

 

0

 

0

 

0.0

 

1

 

1

 

0.7

 

0

 

0

 

0.0

Xiphoid cartilage - partially split

14

11

9.9

9

8

5.7

8

8

5.1

14

11

10.1

Pectoral Girdle

Scapula - misshapen 

 

1

 

1

 

0.6

 

5

 

5

 

6.6

 

2

 

2

 

1.0

 

4

 

2

 

2.6

Pelvic Girdle

Ischium - not ossified

 

0

 

0

 

0.0

 

0

 

0

 

0.0

 

1

 

1

 

0.7

 

0

 

0

 

0.0

Ischium - incomplete ossification

1

1

0.7

1

1

0.7

4

2

2.6

2

2

1.2

Pubis - not ossified

0

0

0.0

0

0

0.0

3

3

1.8

1

1

0.7

Pubis - incomplete ossification

8

3

5.4

8

5

5.8

11

8

7.0

13

8

9.3

Forelimbs

Metacarpal - not ossified

 

51

 

20

 

31.8

 

41

 

15

 

27.0

 

65

 

19

 

42.2

 

57

 

17

 

37.7

Metacarpal - incomplete ossification

0

0

0.0

1

1

0.7

4

3

2.9

3

3

1.9

Forepaw phalanges - 1 or more - ossified

14

10

10.5

13

9

12.8

10

5

6.5

10

5

7.0

Humerus - incomplete ossification

2

2

1.3

1

1

0.7

2

2

1.7

7

5

5.6

Humerus - hole

1

1

0.6

0

0

0.0

1

1

0.6

0

0

0.0

Hindlimbs

Metatarsal - 1st - ossified

 

0

 

0

 

0.0

 

1

 

1

 

4.2

 

0

 

0

 

0.0

 

0

 

0

 

0.0

Metatarsal - not ossified

0

0

0.0

0

0

0.0

1

1

0.7

0

0

0.0

Metatarsal - incomplete ossification

3

2

2.2

0

0

0.0

5

4

2.9

2

2

1.3

Femur - incomplete ossification

12

7

7.7

2

2

1.4

9

6

5.9

7

6

4.6

Total

132

24

86.6

118

24

81.4

130

24

82.7

126

24

90.2

 

Conclusions:
The oral administration of the test substance to pregnant rats by oral gavage during gestation at dose levels of 30, 75 and 150 mg/kg bw/day, resulted in treatment-related macroscopic abnormalities detected in females treated with 150 and 75 mg/kg bw/day. The macroscopic findings detected were considered to be the result of localized irritation from the test item formulations and are not considered to reflect systemic toxicity. The ‘No Observed Adverse Effect Level’ (NOAEL) for the pregnant female was therefore considered to be 150 mg/kg bw/day.
No treatment-related changes were detected in the offspring parameters measured or on embryofetal development. The ‘No Observed Effect Level’ (NOEL) for developmental toxicity was therefore considered to be 150 mg/kg bw/day.
Effect on developmental toxicity: via oral route
Endpoint conclusion:
no adverse effect observed
Dose descriptor:
NOAEL
150 mg/kg bw/day
Study duration:
subacute
Species:
rat
Effect on developmental toxicity: via inhalation route
Endpoint conclusion:
no study available
Effect on developmental toxicity: via dermal route
Endpoint conclusion:
no study available
Additional information

A pre-natal development toxicity study via oral gavage is performed in female Sprague-Dawley rats (OECD Guideline 414). The test item was administered by gavage to three groups each of twenty-four time mated Sprague-Dawley Crl:CD®(SD) IGS BR strain rats, between Days 3 and 19 of gestation inclusive at dose levels 30, 75, and 150 mg/kg bw/day. A further group of twenty-four time mated females was exposed to the vehicle only (Arachis Oil) to serve as a control.

Clinical signs, body weight change, food and water consumptions were monitored during the study.

All females were terminated on Day 20 of gestation and subjected to gross necropsy including examination of the uterine contents. The number of corpora lutea, number, position and type of implantation, placental weights, fetal weight, sex and external and internal macroscopic appearance were recorded. Half of each litter were examined for detailed skeletal development and the remaining half were subjected to detailed visceral examination.

There were no unscheduled deaths. Instances of increased salivation were evident in all females treated with 150 mg/kg bw/day between Days 13 and 19 and in two females treated with 75 mg/kg bw/day on one occasion. No such effects were detected in females treated with 30 mg/kg bw/day. No adverse effect was detected in body weight development and on food and water consumption. Twenty-one females treated with 150 mg/kg bw/day and eleven females treated with 75 mg/kg bw/day had ulcerated stomachs at necropsy. One female treated with 150 mg/kg bw/day also had a pale stomach. One female treated with 30 mg/kg bw/day had white ridges on the non-glandular region of the stomach.

The number of implantations, subsequent embryofetal survival, live litter size and sex ratio on Day 20 of gestation were considered to be unaffected by maternal treatment at 30, 75 or 150 mg/kg bw/day. Mean fetal, placental and litter weights were also considered to have been unaffected by maternal treatment at 30, 75 or 150 mg/kg bw/day.

External examination of fetuses on Day 20 of gestation did not indicate any obvious effect of maternal treatment on fetal development at 30, 75 or 150 mg/kg bw/day. Findings at detailed skeletal and visceral examinations of fetuses on Day 20 of gestation did not indicate any obvious effect of maternal treatment on fetal development at 30, 75 or 150 mg/kg bw/day.

The oral administration of the test subtance to pregnant rats by oral gavage during gestation at dose levels of 30, 75 and 150 mg/kg bw/day, resulted in treatment-related macroscopic abnormalities detected in females treated with 150 and 75 mg/kg bw/day. The macroscopic findings detected were considered to be the result of localized irritation from the test item formulations and are not considered to reflect systemic toxicity.

The ‘No Observed Adverse Effect Level’ (NOAEL) for the pregnant female was therefore considered to be 150 mg/kg bw/day.

No treatment-related changes were detected in the offspring parameters measured or on embryofetal development. The ‘No Observed Effect Level’ (NOEL) for developmental toxicity was therefore considered to be 150 mg/kg bw/day.

Justification for classification or non-classification

Based on the available data and according to the CLP criteria, the test substance should not be classified as toxic to reproduction.