Registration Dossier

Administrative data

Key value for chemical safety assessment

Effects on fertility

Description of key information

Guideline (OECD 421 and 422) reproductive toxicity screening studies have been conducted in rats, for nine members of the higher olefin category, covering C6 to C18. These investigations have all used oral (gavage) exposure. Oral toxicity data is supported by results from a 90-day sub-chronic inhalation study in rats with a C8 olefin. 

 

Except for one study with Nonene, branched, where the NOAEL for reproductive toxicity was considered to be 300 mg/Kg bw/day based on reduced post-natal offspring viability, offspring body weight gain, and litter size at 1000 mg/Kg bw/day, no effects on reproduction were observed in any other study at oral dose levels up to 1000 mg/Kg bw/day.

 

Inhalation exposure up to 10,326 mg/m3 did not reveal any effects on reproductive organs.

Link to relevant study records
Reference
Endpoint:
screening for reproductive / developmental toxicity
Type of information:
experimental study
Adequacy of study:
key study
Study period:
22 August 2013 - 17 October 2013
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: The study is GLP compliant and follow the OECD 422 guideline. Acceptable with no restrictions.
Justification for type of information:
A discussion and report on the read across strategy is given as an attachment in Section 13.
Reason / purpose:
read-across: supporting information
Qualifier:
according to
Guideline:
OECD Guideline 422 (Combined Repeated Dose Toxicity Study with the Reproduction / Developmental Toxicity Screening Test)
Deviations:
no
GLP compliance:
yes (incl. certificate)
Specific details on test material used for the study:
- Name of test material: Nonene, branched
- CAS No.:97280-95-0
- Substance type: UVCB
- Physical state: Clear colorless liquid
- Analytical purity: 100%
- Lot/batch No.: TQ-12-2003-23042013
- Data received: 29 April 2013
- Label: Nonene Sampling Date Apr 22.2013
- Expiration date of the lot/batch: 23 April2014

Nonene, branched, CAS# 97280-95-0 used in this study was a typical production sample, according to the details included in the boundary composition in IUCLID section 1.2. Nonene, branched was chosen in the Higher Olefins category testing strategy because it represents a substance with high tri-sub content (category range 1 – 65%), odd carbon number (C9 odd 90%). Please see the testing strategy attached in section 13 for further details.
Species:
rat
Strain:
other: Wistar Han™:RccHan™:WIST strain rats
Sex:
male/female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Harlan Laboratories U.K. Ltd., Blackthorn, Bicester, Oxon, UK.
- Age at study initiation: 12 weeks old
- Weight at study initiation: (P) Males: 311 to 375 g; Females: 194 to 222 g.
- Housing: Pre-mating: groups of four animals were housed in solid floor polypropylene cages with stainless steel mesh lids and softwood flake bedding (Datesand Ltd., Cheshire, UK). Pairing phase: polypropylene grid floor cages suspended over trays lined with absorbent paper on a one male: one female basis within each dose group. Post mating: the males were returned to their original cages. Mated females were housed individually during gestation and lactation in solid floor polypropylene cages with stainless steel mesh lids and softwood flakes.
- Use of restrainers for preventing ingestion (if dermal): yes/no
- Diet (e.g. ad libitum): A pelleted diet (Rodent 2018C Teklad Global Certified Diet, Harlan Laboratories U.K. Ltd., Oxon, UK.)
- Water (e.g. ad libitum): free access to food and water
- Acclimation period: 5 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 21 +/-2°C
- Humidity (%): 55 =/- 15%
- Air changes (per hr): at least 15 air changes per hour. The low intensity fluorescent lighting was controlled to give twelve hours continuous light
- Photoperiod (hrs dark / hrs light): 12 hr dark and 12 hr light

The study was performed between 10 June 2013 and 10 June 2014. The in-life phase of the study was conducted between 22 August 2013 (first day of treatment) and 17 October 2013 (final day of necropsy).



Route of administration:
oral: gavage
Vehicle:
arachis oil
Details on exposure:
PREPARATION OF DOSING SOLUTIONS:

DIET PREPARATION
- Mixing appropriate amounts with (Type of food): Arachis oil BP
- Storage temperature of food: 4 C

VEHICLE
- Amount of vehicle (if gavage): 4 ml/kg
- Concentration in vehicle:
- Control: 0 ml/kg
- Low: 25 mg/ml
- Intermediate: 75 mg/ml
- High: 250 mg/ml
- Lot/batch no. (if required): 2018C-020713MA
- The volume of the test and control item administrated to each animal was based on the most recent scheduled body weight and was adjusted at weekly intervals.
Details on mating procedure:
- M/F ratio per cage: 1/1
- Length of cohabitation:15 days
- Proof of pregnancy referred to as day 0 of pregnancy
- After successful mating each pregnant female was caged individually during gestation and lactation in solid floor polypropylene cages.
Analytical verification of doses or concentrations:
yes
Duration of treatment / exposure:
6 weeks
Frequency of treatment:
The test item was administrated daily by gavage using a stainless cannula attached to a disposable plastic syringe. Control animals were treated in an identical manner with 4 ml/kg of Arachis oil BP.
Details on study schedule:
i. Groups of twelve male and twelve female animals were treated daily at the appropriate dose level throughout the study (except for females during parturition where applicable). The first day of dosing was designated as Day 1 of the study.
ii. Prior to the start of treatment and once weekly thereafter, all animals were observed for signs of functional/behavioral toxicity.
iii. On Day 15, animals were paired on a 1 male: 1 female basis within each dose group for a maximum of fourteen days.
iv. Following evidence of mating (designated as Day 0 post coitum) the males were returned to their original cages and females were transferred to individual cages.
v. On completion of the pairing phase (during Week 6), five selected males per dose group were evaluated for functional/sensory responses to various stimuli.
vi. Pregnant females were allowed to give birth and maintain their offspring until Day 5 post partum. Litter size, offspring weight and sex, surface righting and clinical signs were also recorded during this period.
vii. At Day 4 post partum, five selected females per dose group were evaluated for functional/sensory responses to various stimuli.
viii. Blood samples were taken from five males from each dose group for hematological and blood chemical assessments on Day 42. The male dose groups were killed and examined macroscopically on Day 43.
ix. Blood samples were taken from five randomly selected females from each dose group for hematological and blood chemical assessment on Day 4 post partum. At Day 5 post partum, all females and surviving offspring were killed and examined macroscopically. Any female which did not produce a pregnancy was also killed and examined macroscopically.
Dose / conc.:
100 mg/kg bw/day (actual dose received)
Remarks:
Doses / Concentrations:

Basis: actual ingested 100 mg/kg bw/day
Dose / conc.:
300 mg/kg bw/day (actual dose received)
Remarks:
Doses / Concentrations:

Basis: actual ingested 300 mg/kg bw/day
Dose / conc.:
1 000 mg/kg bw/day (actual dose received)
Remarks:
Doses / Concentrations:

Basis: actual ingested 1000 mg/kg bw/day
No. of animals per sex per dose:
Three groups, each of twelve males and twelve females
Control animals:
yes, concurrent vehicle
Details on study design:
- Dose selection rationale: Dose selected during the Range-finding experiment - Study Number 41301414

Parental animals: Observations and examinations:
Clinical Observations:
Signs of toxicity, ill-health and behavioural change immediately before dosing, soon after dosing, and one hour after dosing throughout the treatment period.

Functional Observations:
All animals were observed for signs of functional/behavioural toxicity prior to the start of treatment and at weekly intervals thereafter.
Functional performance tests were also performed on five selected males and females from each dose level, prior to termination, together with an assessment of sensory reactivity to various stimuli.
- behavioural assessment
- functional Performance Tests
- Sensory Reactivity

Body Weight:
Individual body weights: Day 1 (prior to dosing), weekly for surviving males until termination and weekly for females until mating was evident.
Female: Days 0, 7, 14 and 20 post coitum, and on Days 1, 4 post partum and terminal kill.

Food Consumption:
Pre-pairing period and after (male): weekly
Females showing evidence of mating: post coitum Days 0-7, 7-14 and 14-20.
Females with live litters: Days 1 and 4 post partum.

Food efficiency:
Males (throughout the study period-with the exception of the mating phase, and females during the pre-pairing phase

Water Consumption:
Daily- visual inspection of the water bottles

Reproductive Performance:
Presence of ejected copulation plugs and each female was examined for the presence of a copulation plug in the vagina.
A vaginal smear was prepared for each female and the stage of oestrus or the presence of sperm was recorded.
The presence of sperm within the vaginal smear and/or vaginal plug in situ was taken as positive evidence of mating (Day 0 of gestation).

Pregnancy and Parturition:
Pregnant females: observations at approximately 0830, 1230 and 1630 hours and around the period of expected parturition (08.30 and 12.30 hr at weekends and public holidays)

Female data collections:

i. Date of pairing
ii. Date of mating
iii. Date and time of observed start of parturition
iv. Date and time of observed completion of parturition

Laboratory Investigations:
- Haematology
- Blood Chemistry
Litter observations:
Number of live and dead offspring
For each litter the following was recorded:
i. Number of offspring born
ii.Number of offspring alive recorded daily and reported on Days 1 and 4 post partum
iii. Sex of offspring on Days 1 and 4 post partum
iv. Clinical condition of offspring from birth to Day 5 post partum
v. Individual offspring weights on Days 1 and 4 post partum (litter weights were calculated retrospectively from this data)

All live offspring were assessed for surface righting reflex on Day 1 post partum.
Postmortem examinations (parental animals):
Pathology Females: Uterus (for signs of implantation and the number of uterine implantations in each horn), the corpora lutea. All adult animals and offspring: full external and internal examination. All macroscopic abnormalities were recorded.
- Organ Weights- Histopathology: Tissues from 5 selected male and female animals/group were preserved. Histopathological examination was undertaken on tissues from control and high dost animals.
To clarify possible treatment-related changes, histopathological examination was extended to include similarly prepared sections of the liver, kidneys (males only), stomach, thyroid and pituitary from animals in the low and intermediate groups. In addition, male kidney tissue was subject to immunohistochemical examination to confirm the presence of alph-2-microglobulin.
Postmortem examinations (offspring):
All adult animals and offspring, including those dying during the study, were subjected to a full external and internal examination, and any macroscopic abnormalities were recorded.
Statistics:
Please see "Any other information on materials and methods incl. tables" for information on statistics.
Reproductive indices:
- Pre-coital Interval
- Fertility Indices
- Gestation length
- Parturition Index

Offspring viability indices:
- Implantation Losses (%)
- Live Birth and Viability Indices
- Sex Ratio (% males)
Clinical signs:
effects observed, treatment-related
Description (incidence and severity):
1000 and 300 mg/kg bw/day: Increasing salivation in males and females. No effects in animals of either sex treated with 100 mg/kg bw/day.
Mortality:
no mortality observed
Description (incidence):
There were no unscheduled deaths.
Body weight and weight changes:
effects observed, treatment-related
Description (incidence and severity):
Males treated with 1000 mg/kg bw/day showed a statistically significant reduction in body weight gain during Weeks 1 and 3 of treatment. Overall body weight gain was reduced for these males.

No such effects were detected in treated females or in males treated with 300 or 100 mg/kg bw/day.

Females treated with 300 mg/kg bw/day showed a statistically significant increase in cumulative body weight gain during the final week of gestation. An increase in body weight gain is considered not to represent an adverse effect oftreatment therefore the intergroup difference was considered not to be of toxicological importance.
Food consumption and compound intake (if feeding study):
effects observed, treatment-related
Description (incidence and severity):
No adverse effect on food consumption was detected in treated animals when compared to control animals.

Females treated with 1000 mg/kg bw/day showed a statistically significant increase in food consumption during the second week of gestation. An increase in food consumption is considered not to represent an adverse effect of treatment therefore the intergroup difference was considered not to be of toxicological importance.
Food efficiency:
no effects observed
Description (incidence and severity):
No adverse effect on food efficiency was detected in treated animals when compared to control animals.
Water consumption and compound intake (if drinking water study):
effects observed, treatment-related
Description (incidence and severity):
Daily measurements of water bottles revealed a marked increase in water consumption in males treated with 1000 mg/kg bw/day and a slight increase in males treated with 300 or 100 mg/kg bw/day throughout the treatment period. Females from all treatment groups also showed an increase in water consumption throughout the treatment period with statistical significance being achieved throughout gestation and lactation at 1000 mg/kg bw/day.
Haematological findings:
effects observed, treatment-related
Description (incidence and severity):
Males treated with 1000 mg/kg bw/day showed a reduction in hemoglobin. Three out of the five individual values were outside of the normal background range for this parameter.

No toxicologically significant effects were detected in females treated with 1000 mg/kg bw/day or animals of either sex treated with 300 or 100 mg/kg bw/day.

Males treated with 1000 mg/kg bw/day showed a reduction in mean corpuscular hemoglobin. Males treated with 1000 and 300 mg/kg bw/day showed a reduction in mean corpuscular hemoglobin concentration. Males from all treatment groups also showed a reduction in
prothrombin time. All of the individual values were within the normal background range for these parameters, therefore the intergroup differences were considered not to be of toxicological importance. Females from all treatment groups showed a statistically significant increase in neutrophil count. The majority of individual values were within the normal background range for these parameters and a true dose related response was not evident, therefore the intergroup differences were considered not to be of toxicological significance.
Clinical biochemistry findings:
effects observed, treatment-related
Description (incidence and severity):
No toxicologically significant effects were detected in the blood chemical parameters examined.

Males treated with 1000 and 300 mg/kg bw/day showed a statistically significant increase in calcium concentration and a statistically significant reduction in alkaline phosphatase. Males treated with 1000 mg/kg bw/day also showed a reduction in cholesterol. All of the individual values were within the normal ranges for rats ofthe strain and age used and in the absence of a true dose related response or any associated histology correlates the intergroup differences were considered not to be of toxicological importance.
Behaviour (functional findings):
effects observed, treatment-related
Description (incidence and severity):
Behavioral Assessments
Weekly open field arena observations did not reveal any treatment-related effects for treated animals when compared to controls.

Functional Performance Tests
There were no toxicologically significant changes in functional performance. Males from all treatment groups showed a statistically significant increase in fore limb grip strength. The statistically significant difference was confined to one out of the three tests and in the absence of a true dose related response or any supporting clinical observations to suggest an effect of neurotoxicity, the intergroup differences were considered to be of no toxicological significance.

Sensory Reactivity Assessments
There were no treatment-related changes in sensory reactivity.
Organ weight findings including organ / body weight ratios:
effects observed, treatment-related
Description (incidence and severity):
Animals of either sex treated with 1000 and 300 mg/kg bw/day and females treated with 100 mg/kg bw/day showed an increase in absolute and relative liver weight. Males treated with 1000 mg/kg bw/day also showed an increase in kidney weight both absolute and relative to terminal body weight. Males from all treatment groups showed an increase in absolute and relative thyroid weight whilst females treated with 1000 mg/kg bw/day showed a reduction in absolute and relative thyroid weight. All of the individual values were within the normal range for rats of the strain and age used and the effect on male thyroid weight also did not show a true dose related response. However with the associated microscopic thyroid changes observed in this study the intergroup differences detected in thyroid weights can not be excluded as an effect of treatment.

Females treated with 1000 mg/kg bw/day showed a reduction in pituitary weight and an increase in kidney weight both absolute and relative to terminal body weight. In the absence of any associated histopathological changes detected in the kidneys or pituitary of females the intergroup differences were considered not to be of toxicological importance.
Gross pathological findings:
effects observed, treatment-related
Description (incidence and severity):
Nine males treated with 1000 mg/kg bw/day had enlarged kidneys at necropsy, eight of which had pale kidneys; four showed a mottled appearance to the kidneys and five of these males also had a dark liver at necropsy. One male treated with 300 mg/kg bw/day had enlarged kidneys at necropsy.

No toxicologically significant effects were detected in females from any treatment group or in males treated with 100 mg/kg bw/day.

One male treated with 100 mg/kg bw/day had small testes and epididymides. One female treated with 300 mg/kg bw/day and two females treated with 100 mg/kg bw/day had reddened lungs at necropsy. Histopathological examination of these tissues did reveal associated microscopic findings to the macroscopic abnormalities however in the absence of a similar effect seen at 1000 mg/kg bw/day, the intergroup differences were considered to be incidental and of no toxicological importance.
Histopathological findings: non-neoplastic:
effects observed, treatment-related
Description (incidence and severity):
The following treatment related microscopic findings were detected:

Liver:hepatocyte hypertrophy was evident in animals of either sex treated with 1000 and 300 mg/kg bw/day and in males treated with 100 mg/kg bw/day. No such effects were detected in females treated with 100 mg/kg bw/day.

Kidneys:proximal tubular basophilia and hyaline droplets were evident in males from all treatment groups. Tubular degeneration/debris was evident in males treated with 1000 and 300 mg/kg bw/day. These findings were demonstrated by immunohistochemical staining to be
due to alpha-2-microglobulin nephropathy. No such effects were detected in any treated female.

Stomach:epithelial hyperplasia was evident in the fore stomach of animals of either sex treated with 1000 mg/kg bw/day and in males treated with 300 mg/kg bw/day. No such effects were detected in females treated with 300 mg/kg bw/day or animals of either sex treated with 100 mg/kg bw/day.

Thyroid:follicular cell hypertrophy and hyperplasia was evident in animals of either sex treated with 1000 and 300 mg/kg bw/day and in males treated with 100 mg/kg bw/day. No such effects were detected in females treated with 100 mg/kg bw/day.

Pituitary:increased incidence and severity of hypertrophic/vacuolated cells in pars anterior was evident in males treated with 1000 and 300 mg/kg bw/day. No such effects were detected in any treated female or in males treated with 100 mg/kg bw/day.
Other effects:
no effects observed
Reproductive function: oestrous cycle:
no effects observed
Reproductive function: sperm measures:
no effects observed
Reproductive performance:
effects observed, non-treatment-related
Description (incidence and severity):
Mating
There were no treatment-related effects on mating performance.

Fertility
No treatment-related effects on fertility were detected for treated animals, when compared to controls.

One control female and one female treated 100 mg/kg bw/day did not achieve pregnancy following evidence of mating. No histopathological correlates were evident in the female or male reproductive organs which could have been the cause of the infertility. One female treated with 1000 mg/kg bw/day failed to show any positive signs of mating and was non pregnant.

Gestation Length
There were no differences in gestation lengths. The distribution for treated females was comparable to controls. All animals showed gestation lengths of 22 to 24½ days.
Clinical Signs:
1000 and 300 mg/kg bw/day: Increasing salivation in males and females. No effects in animals of either sex treated with 100 mg/kg bw/day.

Body Weight and Weight Changes:
Males treated with 1000 mg/kg bw/day showed a statistically significant reduction in body weight gain during Weeks 1 and 3 of treatment. Overall body weight gain was reduced for these males.

No such effects were detected in treated females or in males treated with 300 or 100 mg/kg bw/day.

Females treated with 300 mg/kg bw/day showed a statistically significant increase in cumulative body weight gain during the final week of gestation. An increase in body weight gain is considered not to represent an adverse effect oftreatment therefore the intergroup difference was considered not to be of toxicological importance.

Food consumption and Compound Intake:
No adverse effect on food consumption was detected in treated animals when compared to control animals.

Females treated with 1000 mg/kg bw/day showed a statistically significant increase in food consumption during the second week of gestation. An increase in food consumption is considered not to represent an adverse effect of treatment therefore the intergroup difference was considered not to be of toxicological importance.

Water Consumption:
Daily measurements of water bottles revealed a marked increase in water consumption in males treated with 1000 mg/kg bw/day and a slight increase in males treated with 300 or 100 mg/kg bw/day throughout the treatment period. Females from all treatment groups also showed an increase in water consumption throughout the treatment period with statistical significance being achieved throughout gestation and lactation at 1000 mg/kg bw/day.

Hematology:
Males treated with 1000 mg/kg bw/day showed a reduction in hemoglobin. Three out of the five individual values were outside of the normal background range for this parameter.

No toxicologically significant effects were detected in females treated with 1000 mg/kg bw/day or animals of either sex treated with 300 or 100 mg/kg bw/day.

Males treated with 1000 mg/kg bw/day showed a reduction in mean corpuscular hemoglobin. Males treated with 1000 and 300 mg/kg bw/day showed a reduction in mean corpuscular hemoglobin concentration. Males from all treatment groups also showed a reduction in
prothrombin time. All of the individual values were within the normal background range for these parameters, therefore the intergroup differences were considered not to be of toxicological importance. Females from all treatment groups showed a statistically significant increase in neutrophil count. The majority of individual values were within the normal background range for these parameters and a true dose related response was not evident, therefore the intergroup differences were considered not to be of toxicological significance.

Clinical Biochemistry:
No toxicologically significant effects were detected in the blood chemical parameters examined.

Males treated with 1000 and 300 mg/kg bw/day showed a statistically significant increase in calcium concentration and a statistically significant reduction in alkaline phosphatase. Males treated with 1000 mg/kg bw/day also showed a reduction in cholesterol. All of the individual values were within the normal ranges for rats ofthe strain and age used and in the absence of a true dose related response or any associated histology correlates the intergroup differences were considered not to be of toxicological importance.

Behaviour:
Functional Performance Tests - There were no toxicologically significant changes in functional performance. Males from all treatment groups showed a statistically significant increase in fore limb grip strength. The statistically significant difference was confined to one out of the three tests and in the absence of a true dose related response or any supporting clinical observations to suggest an effect of neurotoxicity, the intergroup differences were considered to be of no toxicological significance.

Organ Weight:
Animals of either sex treated with 1000 and 300 mg/kg bw/day and females treated with 100 mg/kg bw/day showed an increase in absolute and relative liver weight. Males treated with 1000 mg/kg bw/day also showed an increase in kidney weight both absolute and relative to terminal body weight. Males from all treatment groups showed an increase in absolute and relative thyroid weight whilst females treated with 1000 mg/kg bw/day showed a reduction in absolute and relative thyroid weight. All of the individual values were within the normal range for rats of the strain and age used and the effect on male thyroid weight also did not show a true dose related response. However with the associated microscopic thyroid changes observed in this study the intergroup differences detected in thyroid weights can not be excluded as an effect of treatment.

Females treated with 1000 mg/kg bw/day showed a reduction in pituitary weight and an increase in kidney weight both absolute and relative to terminal body weight. In the absence of any associated histopathological changes detected in the kidneys or pituitary of females the intergroup differences were considered not to be of toxicological importance.

Necropsy:
Nine males treated with 1000 mg/kg bw/day had enlarged kidneys at necropsy, eight of which had pale kidneys; four showed a mottled appearance to the kidneys and five of these males also had a dark liver at necropsy. One male treated with 300 mg/kg bw/day had enlarged kidneys at necropsy.

No toxicologically significant effects were detected in females from any treatment group or in males treated with 100 mg/kg bw/day.

One male treated with 100 mg/kg bw/day had small testes and epididymides. One female treated with 300 mg/kg bw/day and two females treated with 100 mg/kg bw/day had reddened lungs at necropsy. Histopathological examination of these tissues did reveal associated microscopic findings to the macroscopic abnormalities however in the absence of a similar effect seen at 1000 mg/kg bw/day, the intergroup differences were considered to be incidental and of no toxicological importance.

Histopathology:
Liver:hepatocyte hypertrophy was evident in animals of either sex treated with 1000 and 300 mg/kg bw/day and in males treated with 100 mg/kg bw/day. No such effects were detected in females treated with 100 mg/kg bw/day.

Kidneys:proximal tubular basophilia and hyaline droplets were evident in males from all treatment groups. Tubular degeneration/debris was evident in males treated with 1000 and 300 mg/kg bw/day. These findings were demonstrated by immunohistochemical staining to be
due to alpha-2-microglobulin nephropathy. No such effects were detected in any treated female.

Stomach:epithelial hyperplasia was evident in the fore stomach of animals of either sex treated with 1000 mg/kg bw/day and in males treated with 300 mg/kg bw/day. No such effects were detected in females treated with 300 mg/kg bw/day or animals of either sex treated with 100 mg/kg bw/day.

Thyroid:follicular cell hypertrophy and hyperplasia was evident in animals of either sex treated with 1000 and 300 mg/kg bw/day and in males treated with 100 mg/kg bw/day. No such effects were detected in females treated with 100 mg/kg bw/day.

Pituitary:increased incidence and severity of hypertrophic/vacuolated cells in pars anterior was evident in males treated with 1000 and 300 mg/kg bw/day. No such effects were detected in any treated female or in males treated with 100 mg/kg bw/day.

Reproductive Performance:
Fertility - No treatment-related effects on fertility were detected for treated animals, when compared to controls.

One control female and one female treated 100 mg/kg bw/day did not achieve pregnancy following evidence of mating. No histopathological correlates were evident in the female or male reproductive organs which could have been the cause of the infertility. One female treated with 1000 mg/kg bw/day failed to show any positive signs of mating and was non pregnant.
Key result
Dose descriptor:
NOAEL
Effect level:
1 000 mg/kg bw/day (actual dose received)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
other: Systemic Toxicity
Dose descriptor:
NOEL
Effect level:
300 mg/kg bw/day (actual dose received)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
other: Reproductive Toxicity
Key result
Critical effects observed:
no
Clinical signs:
effects observed, non-treatment-related
Description (incidence and severity):
No obvious clinical signs of toxicity were detected for offspring from treated females when compared to controls. The incidental clinical signs detected throughout the control and treated groups, consisting of small size, cold, weak, pale, no milk in stomach, physical injury, found dead or missing, were considered to be low incidence findings observed in offspring in studies of this type and were considered unrelated to test item toxicity.
Mortality / viability:
no mortality observed
Body weight and weight changes:
effects observed, treatment-related
Description (incidence and severity):
Litter weights on Day 4 post partum from females treated with 1000 mg/kg bw/day were reduced when compared to control litters however statistical significance was not achieved. Offspring body weight gain at 1000 mg/kg bw/day was also reduced between Days 1 and 4 post partum. Statistical significance was achieved for male off spring only. No such effects were detected in litters from females treated with 300 or 100 mg/kg bw/day.
Sexual maturation:
not examined
Organ weight findings including organ / body weight ratios:
no effects observed
Gross pathological findings:
effects observed, non-treatment-related
Description (incidence and severity):
No treatment-related macroscopic abnormalities were detected for interim death or terminal kill offspring. The incidental findings observed were those occasionally observed in reproductive studies of this type and were considered to be unrelated to toxicity of the test item.
Histopathological findings:
no effects observed
Behaviour (functional findings):
no effects observed
Description (incidence and severity):
Statistical analysis of surface righting reflex data did not reveal any significant intergroup differences.
Offspring Litter Size, Sex Ratio and Viability
No significant differences were detected for corpora lutea, implantation counts or implantation losses for treated animals when compared to controls. Statistical analysis of the data did not reveal any significant intergroup differences.

Of the litters born, litter size at birth and subsequently on Day 1 post partumwere comparable to controls. Litter size on Day 4 post partumhowever was reduced in litters from females treated with 1000 mg/kg bw/day although statistical significance was not achieved. Offspring viability on Day 4 post partumwas statistically significantly reduced in these litters. A total litter loss was also observed for one female treated with 1000 mg/kg bw/day. No such effects were detected in litters from females treated with 300 or 100 mg/kg bw/day.

There were no intergroup differences in sex ratio (percentage male offspring) for litters from treated groups compared to controls. Statistical analysis of the data did not reveal any significant intergroup differences.

Offspring Growth and Development
Litter weights on Day 4 post partumfrom females treated with 1000 mg/kg bw/day were reduced when compared to control litters however statistical significance was not achieved. Offspring body weight gain at 1000 mg/kg bw/day was also reduced between Days 1 and 4 post partum. Statistical significance was achieved for male offspring only.

No such effects were detected in litters from females treated with 300 or 100 mg/kg bw/day. Statistical analysis of surface righting reflex data did not reveal any significant intergroup differences.

No obvious clinical signs of toxicity were detected for offspring from treated females when compared to controls. The incidental clinical signs detected throughout the control and treated groups, consisting of small size, cold, weak, pale, no milk in stomach, physical injury, found dead or missing, were considered to be low incidence findings observed in offspring in studies of this type and were considered unrelated to test item toxicity.
Key result
Dose descriptor:
NOEL
Generation:
F1
Effect level:
300 mg/kg bw/day (actual dose received)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
other: reduced post natal offspring viability, offspring body weight gain and litter size at 1000 mg/kg bw/day.
Key result
Critical effects observed:
no
Key result
Reproductive effects observed:
yes
Lowest effective dose / conc.:
1 000 mg/kg bw/day (actual dose received)
Treatment related:
yes
Relation to other toxic effects:
reproductive effects occurring together with other toxic effects, but not as a secondary non-specific consequence of other toxic effects
Dose response relationship:
not specified
Relevant for humans:
not specified

Table 2. Summary of Reproductive Performance - Groups Values

 

Dose Group (mg/Kg bw/day

0 (Control)

100

300

1000

Males

 

Initial group size

12

12

12

12

Paired

12

12

12

12

Failed to mate

0

0

0

1

Failed to induce pregnancy in female partner

1

1

0

0

Induced pregnancy in female partner

11

11

12

11

Surviving to terminal necropsy

12

12

12

12

 

Females

 

Initial group size

12

12

12

12

Paired

12

12

12

12

Failed to mate

0

0

0

1

Non-pregnant

1

1

0

0

Rearing young to Day 5 of age

11

11

12

10

Total Litter Loss

0

0

0

1

 

Table 3. Group Mean Functional Test Values and Standard Deviations - Males

Group

(sex)

 

Test 1 Forelimb

(g)

Test 1 Hindlimb

(g)

Test 2 Forelimb

(g)

Test 2 Hindlimb

(g)

Test 3 Forelimb

(g)

Test 3 Hindlimb

(g)

Overall

Activity

Overall Mobile

Last 20%

Activity

Last 20% Mobile

1 (M)

Mean

1143.2

422.6

1004.2

524.4

985.2

445.0

377.2

0.6

3.4

0.0n

S.D.

234.7

131.5

113.9

66.2

91.6

76.7

122.9

0.9

4.0

0.0

N

5

5

5

5

5

5

5

5

5

5

 

2 (M)

Mean

1176.8

510.2

1303.0*

581.4

1084.8

628.2

384.0

0.6

8.2

0.0n

S.D.

237.0

140.4

313.6

105.2

146.7

157.4

214.5

0.5

9.5

0.0

N

5

5

5

5

5

5

5

5

5

5

 

3 (M)

Mean

1353.0

593.8

1450.4*

472.0

1378.2

528.2

334.6

0.2

4.2

0.0n

S.D.

297.1

121.5

200.0

147.6

442.9

177.5

141.7

0.4

5.2

0.0

N

5

5

5

5

5

5

5

5

5

5

 

4 (M)

Mean

1071.2

620.8

1271.0*

473.4

1118.6

555.0

250.8

0.2

36.2

0.0n

S.D.

257.2

184.4

250.8

230.9

185.2

151.8

118.6

0.4

58.3

0.0

N

5

5

5

5

5

5

5

5

5

5

General Footnote: Unit = Time (seconds) for Motor Activity Assessments

Dose Levels: Group 1 - 0(Control); Group 2 - 100 mg/kg bw/day; Group 3 - 300 mg/kg bw/day; Group 4 - 1000 mg/kg bw/day

* Significantly different from control group p<0.05

n Data not appropriate for statistical analysis

 

Table 4. Group Mean Body Weight Values - Males

 

Group

(sex)

 

Increase in Body Weight (g)

 

Day Numbers Relative to Start Date

Abs Gain

% Gain

From:

1

8

15

22

29

36

1

1

To:

8

15

22

29

36

43

43

43

1 (M)

Mean

14.4

13.0

9.7

14.6

10.2

6.8

68.6

20.9

S.D.

5,8

6.5

6.6

6.0

4.5

3.7

21.7

5.9

N

12

12

12

12

12

12

12

12

 

 

 

2 (M)

Mean

13.9

12.5

6.9

13.3

12.0

4.3

63.0

19.0

S.D.

3.8

3.9

6.3

4.4

3.4

5.1

16.9

4.8

N

12

12

12

12

12

12

12

12

 

 

 

3 (M)

Mean

13.4

15.3

7.3

14.1

11.9

8.1

70.2

21.3

S.D.

5.5

5.0

3.9

4.6

5.6

3.4

18.8

5.3

N

12

12

12

12

12

12

12

12

 

 

 

4 (M)

Mean

5.5**

10.0

1.7**

12.4

8.1

5.1

42.8

13.0

S.D.

9.8

7.5

6.8

3.8

4.8

4.8

18.0

5.3

N

12

12

12

12

12

12

12

12

Dose Levels: Group 1 - 0(Control); Group 2 - 100 mg/kg bw/day; Group 3 - 300 mg/kg bw/day; Group 4 - 1000 mg/kg bw/day

** Significantly different from control group p<0.01

 

Table 5. Group Mean Body Weight Gains - Females

 

Group

(sex)

 

Increase in Body Weight (g)

Cumulative Body Weight Change (g)

 

Days

Days

 

Gestation

Lactation

Gestation

From:

0

7

14

1

0

0

To:

7

14

20

4

14

20

1 (F)

Mean

29.8

28.9

60.4

7.9

58.7

119.1

S.D.

4.4

3.9

13.0

5.4

7.3

16.3

N

11

11

11

11

11

11

 

2 (F)

Mean

27.4

28.3

60.8

11.8

55.6

116.5

S.D.

4.8

5.0

12.5

10.3

6.1

14.9

N

11

11

11

11

11

11

 

3 (F)

Mean

33.3

33.1

69.1

12.8

66.4

135.5*

S.D.

5.2

5.2

10.9

7.1

7.8

15.2

N

12

12

12

12

12

12

 

4 (F)

Mean

29.8

27.9

57.3

6.4

57.7

115.0

S.D.

6.8

6.5

11.7

12.2

11.4

17.1

N

11

11

11

10

11

11

Dose Levels: Group 1 - 0(Control); Group 2 - 100 mg/kg bw/day; Group 3 - 300 mg/kg bw/day; Group 4 - 1000 mg/kg bw/day

* Significantly different from control group p<0.05

 

Table 6. Group Mean Food Consumptions - Females

Group

(sex)

 

Day Numbers

Gestation

Lactation

From

0

7

14

1

To

7

14

20

4

1 (F)

Mean

19.7

22.3

25.1

26.5

S.D.

1.8

1.9

2.4

4.6

N

11

11

11

11

 

2 (F)

Mean

20.2

22.5

24.6

29.5

S.D.

2.0

2.3

1.2

8.3

N

11

11

11

11

 

3 (F)

Mean

20.7

23.6

25.5

28.2

S.D.

1.2

1.2

1.1

3.3

N

12

12

12

12

 

4 (F)

Mean

20.9

24.2*

26.8

25.4

S.D.

1.6

2.2

2.3

6.2

N

11

11

11

10

Dose Levels: Group 1 - 0(Control); Group 2 - 100 mg/kg bw/day; Group 3 - 300 mg/kg bw/day; Group 4 - 1000 mg/kg bw/day

* Significantly different from control group p<0.05

 

Table 7. Group Mean Water Consumption - Females

Group

(sex)

 

Day Numbers

Gestation

Lactation

From

0

7

14

1

To

7

14

20

4

1 (F)

Mean

27.6

31.0

36.0

39.9

S.D.

3.8

5.1

6.8

4.3

N

11

11

11

11

 

2 (F)

Mean

32.1

36.9

39.9

44.2

S.D.

5.3

5.9

4.8

10.6

N

11

11

11

11

 

3 (F)

Mean

36.5

42.9

44.8

45.8

S.D.

2.2

13.9

10.7

7.1

N

12

12

12

12

 

4 (F)

Mean

52.6***

67.8***

76.4***

54.1**

S.D.

20.4

22.5

34.1

12.5

N

11

11

11

11

Dose Levels: Group 1 - 0(Control); Group 2 - 100 mg/kg bw/day; Group 3 - 300 mg/kg bw/day; Group 4 - 1000 mg/kg bw/day

** Significantly different from control group p<0.01

*** Significantly different from control group p<0.001

 

Table 8. Group Mean Hematological Values - Males

Group

 

Hb (g/dL)

MCH (pg)

MCHC (g/dL)

CT (seconds)

Group 1 (0 – Control)

Mean

16.92

18.86

35.66

9.92

S.D.

0.44

0.44

1.49

0.47

N

5

5

5

5

 

Group 2 (100 mg/Kg bw/day)

Mean

16.86

18.64

35.18

9.40*

S.D.

0.74

0.11

0.67

0.23

N

5

5

5

5

 

Group 3 (300 mg/Kg bw/day)

Mean

16.08

18.36

34.12**

9.34*

S.D.

0.31

0.48

0.53

0.26

N

5

5

5

5

 

Group 4 (1000 mg/Kg bw/day)

Mean

14.94**

17.58**

33.72**

9.34*

S.D.

1.56

0.63

0.28

0.36

N

5

5

5

5

* Significantly different from control group p<0.05

** Significantly different from control group p<0.01

 

Table 9. Group Mean Hematological Values - Females

Group

 

Neut (109/L)

Group 1 (0 – Control)

Mean

1.360

S.D.

0.417

N

5

 

Group 2 (100 mg/Kg bw/day)

Mean

2.500*

S.D.

0.568

N

5

 

Group 3 (300 mg/Kg bw/day)

Mean

2.146*

S.D.

0.463

N

5

 

Group 4 (1000 mg/Kg bw/day)

Mean

1.872*

S.D.

0.509

N

5

* Significantly different from control group p<0.05

 

Table 10. Group Mean Blood Chemical Values - Males

Group

 

Ca++(mmol/L)

AP (IU/L)

Chol (mg/dL)

Group 1 (0 – Control)

Mean

2.448

234.0

95.2

S.D.

0.258

52.1

5.3

N

5

5

5

 

Group 2 (100 mg/Kg bw/day)

Mean

2.522

201.6

102.6

S.D.

0.146

26.1

14.4

N

5

5

5

 

Group 3 (300 mg/Kg bw/day)

Mean

2.730*

150.6**

99.4

S.D.

0.077

10.8

10.7

N

5

5

5

 

Group 4 (1000 mg/Kg bw/day)

Mean

2.644*

157.6**

78.0*

S.D.

0.129

55.2

13.3

N

5

5

5

* Significantly different from control group p<0.05

** Significantly different from control group p<0.01

 

Table 11. Summary Incidence of Necropsy Findings - Males

 

Males

0

Control

100

mg/Kg bw/day

300

mg/Kg bw/day

1000

mg/Kg bw/day

Number of animals examined

12

12

12

12

Epididymides

 

Submitted

(12)

(12)

(12)

(12)

No Visible Lesions

12

11

12

12

Small; Left

0

1

0

0

 

Kidneys

 

Submitted

(12)

(12)

(12)

(12)

No Visible Lesions

12

12

11

3

Enlarged

0

0

1

9

Mottled Appearance

0

0

0

4

Pallor

0

0

0

3

Pale

0

0

0

6

 

Liver

 

Submitted

(12)

(12)

(12)

(12)

No Visible Lesions

12

12

12

7

Dark

0

0

0

5

 

Testes

 

Submitted

(12)

(12)

(12)

(12)

No Visible Lesions

12

11

12

12

Small; Left

0

1

0

0

 

Table 12. Summary Incidence of Necropsy Findings - Females

 

Females

0

Control

100

mg/Kg bw/day

300

mg/Kg bw/day

1000

mg/Kg bw/day

Number of animals examined

12

12

12

12

Liver

 

Submitted

(12)

(12)

(12)

(12)

No Visible Lesions

12

12

12

11

Dark

0

0

0

1

 

Lungs (With Bronchi)

 

Submitted

(12)

(12)

(12)

(12)

No Visible Lesions

12

10

11

12

Reddened

0

2

1

0

  

Table 13. Group Mean Organ Weights with Corresponding Relative (% of Body Weight) Organ Weights

 

 

Males

Females

0

Control

100

mg/Kg bw/day

300

mg/Kg bw/day

1000

mg/Kg bw/day

0

Control

100

mg/Kg bw/day

300

mg/Kg bw/day

1000

mg/Kg bw/day

Kidneys

Mean (g)

2.29348

2.31240

2.38234

3.16702**

1.54460

1.65510

1.66136

1.97002**

S.D.

0.32697

0.14310

0.16218

0.44018

0.14343

0.11476

0.20712

0.11782

N

5

5

5

5

5

5

5

5

 

Mean (%)

0.574

0.585

0.616

0.830**

0.565

0.610

0.602

0.701**

S.D.

0.046

0.013

0.051

0.102

0.047

0.035

0.055

0.041

N

5

5

5

5

5

5

5

5

 

Liver

Mean (g)

12.7830

13.2647

14.3898**

19.3412**

10.5429

11.7936*

12.7876**

16.1219**

S.D.

1.51792

0.98038

1.33119

1.23327

0.51191

0.52983

1.29755

1.61466

N

5

5

5

5

5

5

5

5

 

Mean (%)

3.203

3.355

3.710**

5.070**

3.857

4.353*

4.638**

5.742**

S.D.

0.125

0.111

0.164

0.230

0.149

0.194

0.424

0.639

N

5

5

5

5

5

5

5

5

 

Pituitary

Mean (g)

 

 

 

 

0.01793

0.01529

0.01783

0.01467**

S.D.

 

 

 

 

0.00277

0.00241

0.00264

0.00180

N

 

 

 

 

11

11

12

10

 

 

Mean (%)

 

 

 

 

0.006

0.006

0.006

0.005**

S.D.

 

 

 

 

0.001

0.001

0.001

0.001

N

 

 

 

 

11

11

12

10

 

Thyroid/Parathyroid

Mean (g)

0.01552

0.02172**

0.02162**

0.02112**

0.002238

0.01972

0.02046

0.01776*

S.D.

0.00377

0.00176

0.00140

0.00207

0.00395

0.00339

0.00202

0.00259

N

5

5

5

5

5

5

5

5

 

Mean (%)

0.004

0.005**

0.006**

0.006**

0.008

0.007

0.007

0.006*

S.D.

0.001

0.000

0.000

0.000

0.002

0.001

0.001

0.001

N

5

5

5

5

5

5

5

5

* Significantly different from control group p<0.05

** Significantly different from control group p<0.01

 

Table 14. Histopathology – Treatment-related Findings

 

Male

Female

Dosage

(mg/Kg bw/day)

0

100

300

1000

0

100

300

1000

 

Liver

Number Examined

5

5

5

9

5

5

5

6

- Hypertrophy; hepatoc.

0

5 (1.0)

5 (1.0)

9 (1.7)

0

0

3 (1.0)

6 (1.0)

- Congestion

0

0

0

5 (1.0)

0

0

0

1(3.0)

 

Thyroid

Number Examined

5

5

5

5

5

5

5

5

- Hypertrophy / Hyperplasia;

Follicular Cell

2 (1.0)

5 (1.4)

5 (1.4)

5 (1.8)

2 (1.0)

2 (1.0)

3 (1.3)

5 (1.2)

 

Pituitary Gland

Number Examined

12

12

12

12

12

12

12

10

-Hypertrophic / Vacuolated cells in

Pars Anterior

10 (1.0)

9 (1.2)

12 (1.4)

9 (1.6)

0

0

3 (1.0)

0

 

Kidney

Number Examined

5

5

5

9

5

5

5

5

- Tubular Basophilia

0

3 (1.0)

4 (1.3)

9 (2.0)

0

0

0

1 (1.0)

- Tubular Degeneration

/ Debris

0

0

4 (1.0)

7 (2.0)

0

0

0

0

- Hyaline Droplets

0

5 (1.0)

5 (1.8)

9 (2.1)

0

0

0

0

 

Kidney Immunohistochemistry

Number Examined

3

3

3

3

0

0

0

0

- alpha-2-microglobulin

3 (1.0)

3 (2.0)

3 (2.3)

3 (2.7)

0

0

0

0

 

Stomach - Forestomach (microscopic changes recorded in the stomach)

Number Examined

5

5

5

5

5

5

5

5

- Hyperplasia, epithelial

0

0

2 (1.0)

2 (1.0)

0

0

0

3 (1.0)

(): Mean Severity

Findings in Bold were considered treatment-related

Table 15. Group Mean offspring Weights

Group

 

Offspring Body Weight Change (g) – Days 1-4

Males

Females

Control (0 mg/Kg bw/day)

Mean (g)

3.18

3.07

S.D.

0.90

0.82

N

10

11

 

100 mg/Kg bw/day

Mean (g)

3.07

3.02

S.D.

0.67

0.61

N

11

11

 

300 mg/Kg bw/day

Mean (g)

2.60

2.54

S.D.

0.62

0.48

N

12

12

 

1000 mg/Kg bw/day

Mean (g)

2.30*

2.42

S.D.

0.85

0.78

N

10

9

* Significantly different from control group p<0.05

 

Table 16. Group Mean Implantation Losses and Survival Indices Values

Group

 

Pre-Implantation

Loss (%)

Post-Implantation

Loss (%)

Live Birth

Index (%)

Viability

Index (%)

Control

(0 mg/Kg bw/day)

Mean

13.9

8.5

100.0

98.9

S.D.

13.7

9.2

0.0

3.8

N

11

11

11

11

 

100 mg/Kg bw/day

Mean

7.9

12.6

100.0

99.2

S.D.

16.2

15.3

0.0

2.5

N

11

11

11

11

 

300 mg/Kg bw/day

Mean

3.4

5.4

100.0

99.4

S.D.

4.9

6.0

0.0

1.9

N

12

12

12

12

 

1000 mg/Kg bw/day

Mean

12.2

12.1

97.1

91.9*

S.D.

18.6

8.2

7.2

9.5

N

11

11

11

10

* Significantly different from control group p<0.05

Conclusions:
Oral administration of Nonene, branched to rats for 8 weeks, at levels of 100, 300 and 1000 mgk/kg/day, resulted in treatment related effects in both sexes and at all treatment levels. The effects seen were however either adaptive in nature or were not considered toxicologically relevant for man. The No Observed Adverse Effect Level (NOAEL) for systemic toxicity was therefore considered to be 1000 mg/kg bw/day.

The ‘No Observed Effect Level’ (NOEL) for reproductive toxicity was considered to be 300 mg/kg bw/day based on reduced offspring viability, offspring body weight gain, litter size and litter weigh at 1000 mg/kg bw/day.
Executive summary:

In a key Guideline (OECD 422) combined repeated dose reproductive/developmental toxicity study, the test material (Nonene, branched; CAS# 97280-95-0) was administered by gavage to three groups of Wistar Han™:RccHan™:WIST strain rats (12/sex/dose) for up to eight weeks (including a two week pre-pairing phase, pairing, gestation and early lactation for females), at dose levels of 100, 300 and 1000 mg/Kg bw/day. A control group of twelve males and twelve females was dosed with vehicle alone (Arachis oil BP).

 

Clinical signs, behavioral assessments, body weight change and food and water consumption were monitored during the study. Pairing of animals within each dose group was undertaken on a one male: one female basis within each treatment group on Day 15 of the study, with females subsequently being allowed to litter and rear their offspring to Day 5 of lactation. During the lactation phase, daily clinical observations were performed on all surviving offspring, together with litter size and offspring weights and assessment of surface righting reflex. Extensive functional observations were performed on five selected males from each dose group after the completion of the pairing phase, and for five selected parental females from each dose group on Day 4 post-partum. Hematology and blood chemistry were evaluated prior to termination on five selected males and females from each dose group.

 

Adult males were terminated on Day 43, followed by the termination of all females and offspring on Day 5 post-partum. Any female which did not produce a pregnancy was terminated on or after Day 25 post coitum. One female which did not show positive evidence of mating and did not produce a pregnancy was terminated on Day 57. All animals were subjected to a gross necropsy examination and histopathological evaluation of selected tissues was performed.

 

No test material related mortality was observed through the study period. Clinical signs were detected in animals of either sex treated with 300 and 1000 mg/Kg bw/day during the study. Episodes of increased salivation were reported from Day 8 onwards. The physical condition of males treated with 1000 mg/Kg bw/day was also affected with reductions in body weight development during Weeks 1 and 3 of treatment. Subsequently, a reduction in overall body weight gain in these males and a slight reduction in food efficiency during week 1, was evident. A reduction in haemoglobin was also observed in these males. Water consumption was also significantly increased for animals of either sex from all treatment groups throughout the treatment period. Observations of this nature are often reported when a test material formulation is unpalatable or irritant and can be associated with gastric irritancy rather than be attributable to systemic toxicity. This was supported microscopically with stomach changes identified as epithelial hyperplasia in the forestomach in animals of either sex treated with 1000 mg/Kg bw/day and in males treated with 300 mg/Kg bw/day. This finding was considered to be the result of local irritancy of the test material and therefore cannot be considered indicative of true systemic toxicity.

 

Although there were some statistically significant differences in treated animals from controls for the blood chemical parameters measured, these differences were considered not to be of toxicological significance. Macroscopic findings detected at necropsy were confined to enlarged, pale and mottled kidneys and a dark liver in a number of males treated with 1000 mg/Kg bw/day. One male treated with 300 mg/Kg bw/day also had enlarged kidneys.

 

Histopathological examination of the liver revealed hepatocellular hypertrophy in animals of either sex treated with 1000 and 300 mg/Kg bw/day and in males treated with 100 mg/Kg bw/day. Organ weight data supported this finding with increased absolute and relative liver weights observed in these animals. In the absence of any degenerative or inflammatory changes, this condition is considered to be adaptive in nature.

 

Microscopic examination of the thyroid revealed increased incidence and severity of follicular cell hypertrophy/hyperplasia in animals of either sex treated with 1000 and 300 mg/Kg bw/day and in males treated with 100 mg/Kg bw/day. Males treated with 1000 and 300 mg/Kg bw/day also showed hypertrophic/vacuolated cells in the pituitary. The thyroid, liver and pituitary changes are characteristic of a consequence of hepatocellular induction as a result of enhanced hepatic metabolism. As a side effect of hepatic induction an increased liver metabolism of thyroid hormones T3 and T4 can occur. This subsequently leads to an enhanced thyroid gland production of these hormones as a consequence of a negative feedback stimulation of TSH production. The appearance of thyroid follicular cell hypertrophy and hypertrophic/vacuolated cells in the pituitary are themselves considered to be a result of this process. The thyroid and pituitary changes were considered to be adaptive in nature.

 

Microscopic examination also revealed effects in the kidneys of males from all treatment groups. Tubular basophilia and hyaline droplets was present in males from all treated groups. These tubular findings were also accompanied by tubular degeneration/debris in males treated with 1000 and 300 mg/Kg bw/day. The hyaline droplets can be directly linked to accumulation of alpha 2u-globulin, which is unique to the male rat. This finding is commonly observed in male rats following treatment with some hydrocarbons and is not predictive of any adverse effect in humans. The remaining kidney findings consisting of tubular degeneration/debris may be considered to represent an adverse effect of the test item, however immunohistochemical staining demonstrated that these findings were correlated to the same condition as hyaline droplets.

 

Mating performance and fertility was unaffected by treatment. Offspring viability was however reduced in litters from females treated with 1000 mg/Kg bw/day on Day 4 post-partum. Subsequently reduced litter size and litter weights were evident in these litters on Day 4 post-partum when compared to controls. One female treated with 1000 mg/Kg bw/day also had a total litter loss between Days 2 and 4 post-partum. The mean offspring body weight gains for litters which survived to Day 5 post-partum were also reduced between Days 1 and 4 post-partum at this treatment level.

 

Based on the results observed, the No Observed Adverse Effect Level (NOAEL) for systemic toxicity was determined to be 1000 mg/Kg bw/day. The ‘No Observed Effect Level’ (NOEL) for reproductive toxicity was considered to be 300 mg/Kg bw/day, based on reduced offspring viability, offspring body weight gain, litter size and litter weights on Day 4 post-partum at 1000 mg/Kg bw/day.

Effect on fertility: via oral route
Endpoint conclusion:
adverse effect observed
Dose descriptor:
NOAEL
300 mg/kg bw/day
Study duration:
subchronic
Species:
rat
Quality of whole database:
In total, nine good quality screening studies are available on nine category members, covering C6 to C18 higher olefins. The study on nonene, branched, in common with all other studies, showed no effects on fertility at levels up tp 1000 mg/Kg. There was however, a slight effect on post-natal pup viability, litter size and weight at 1000 mg/Kg, with a clear NOAEL of 300 mg/Kg. This study was selected as being the 'worst case', although it should be recognized that none of the other eight studies showed any reproductive effects on post-natal pup viability.
Effect on fertility: via inhalation route
Endpoint conclusion:
no study available
Effect on fertility: via dermal route
Endpoint conclusion:
no study available
Additional information

Effects on Fertility

Reproductive toxicity screening studies are available for the following members of the higher olefin Category:

Test substance identity

Testing for Reproductive Effects

Hex-1-ene

OECD 421

Alkenes, C6

OECD 422

Oct-1-ene

OECD 422

Nonene

Branched

OECD 422

Decene

OECD 422

Tetradec-1-ene

OECD 422

Hexadecene

OECD 422

Octadec-1-ene

OECD 422

Octadecene

OECD 421

Information is available from multiple guideline (OECD 421 and 422) studies that investigated the reproductive toxicity potential in rats, for members of the higher olefin category. Studies covered substances in the range C6 to C18, following oral exposure.

C6 members of the category

In a supporting Guideline (OECD 422) combined repeated dose, reproductive/developmental toxicity screening study (Thorsud, 2003), the test material (C6-Olefin/Hexene) was administered to Sprague Dawley CrI:CD®(SD)IGS BR rats (12/sex/dose) once daily (seven days a week) by oral gavage at doses of 0, 100, 500, or 1000 mg/Kg/day for a minimum of 14 days prior to mating and continuing through lactation day 3.

 

Cage-side observations for overt signs of toxicity were performed once daily within approximately one-half hour to two hours following dosing. Detailed clinical observations were performed a minimum of weekly until evidence of mating was detected, daily during gestation and lactation and on the day of scheduled euthanasia. Individual body weights were recorded on days 0, 3, 7 and 12 prior to mating, on gestation days 0, 7, 14 and 20 and on lactation days 1 and 4. Individual food consumption was recorded on the same days as body weights (except during cohabitation). After a minimum of 14 days of treatment, females were cohabited with males from the same treatment group in the toxicity study.

Following parturition, pup viability, sex determinations, observations and body weights were recorded at specified intervals during lactation days 0 and 4. Surviving females and their pups were euthanized and necropsied on lactation day 4. Fresh organ weights were recorded for surviving females and selected tissues and organs were preserved from all females. All gross lesions collected at necropsy from all females were examined microscopically.

 

There were no toxicologically meaningful differences noted in F0 mean body weights, body weight gain or food consumption between the control and test article-treated groups. The F0 female mating and fertility indices and mean gestation lengths were comparable between the control and test article-treated groups. The mean number of F1 pups delivered and the F1 live birth index were comparable between the control and test article-treated groups, and there were no toxicologically meaningful differences in the F1 viability index (pups surviving to lactation day 4) between the groups. The mean F1 pups per litter and the F1 pup sex ratio were comparable between the control and test article-treated groups on lactation days 0 and 4.

 

Mean F1 pup weights were slightly but not statistically lower than controls in the 500 mg/Kg/day group on lactation day 1 and in the 1000 mg/Kg/day group on lactation days 1 and 4. However, the mean body weights in these groups were within the range of SLI historical control data. Mean F1 pup weights in the 100 mg/Kg/day group were comparable to controls on lactation days 1 and 4. There were no toxicologically meaningful differences in pup observations during lactation.

 

No other remarkable findings were noted in the F1 pups during lactation. No remarkable gross necropsy findings were noted for F1 pups found dead during the study or euthanized at study termination. No remarkable internal gross necropsy findings were noted for surviving F0 females in the control or test article-treated groups at study termination. There were no toxicologically meaningful differences in F0 absolute or relative organ weight data between the groups and no toxicologically meaningful microscopic findings were noted for F0 females in the test article-treated groups.

 

Based on the results of this study, a dosage level of 1000 mg/Kg/day was considered a no-observed-adverse-effect level (NOAEL) for reproductive toxicity.

 

In an OECD 421 reproductive/developmental toxicity screening study (Daniel, 1995), 1-hexene was administered via gavage to twelve Sprague-Dawley rats/sex/dose at doses of 0, 100, 500, or 1000 mg/kg/day. The test material was composed of equal amounts of Neodene 6 alpha olefin, hexene-1 Gulftene 6, and alpha olefin C6 1 -hexene, which were obtained from different sources. Males were treated for 44 days beginning 28 days prior to mating and females were treated for 41 to 55 days beginning 14 days prior to mating through lactation day 4.

 

No reproductive or developmental effects were observed. There was a slight, but significant, decrease in absolute epididymal weight at all concentrations. The relative epididymal to brain weight was only significantly decreased in the low-dose group. Although the absolute epididymides weight was significantly decreased in parental males; the change was within 10% of the control, there was no dose response, there was no effects noted microscopically, and there were no effects on fertility. Therefore, this is not considered to be toxicologically significant. Pitted kidneys were observed at necropsy for 2 of 12 mid-dose males and 3 of 12 high-dose males. The predominant microscopic finding in males was the presence of large hyaline droplets in the proximal convoluted tubule that was dose related. These findings suggest hydrocarbon nephropathy, which is a toxicological effect specific to male rats and is not considered relevant to humans.

 

The NOAEL for systemic, reproductive, and developmental toxicity was 1000 mg/kg/day, which excluded the hydrocarbon nephropathy in males.

C8 members of the category

 

A guideline Oral (Gavage) Combined Repeat Dose Toxicity Study with Reproduction/Developmental Toxicity Screening Test in the Rat (OECD 422) was performed on Oct-1-ene (CAS # 111 -66 -0; Harlan Laboratories, 2014).

 

After eight weeks treatment, clinical signs were detected in animals of either sex treated with 1000 mg/Kg bw/day and in males treated with 300 mg/Kg bw/day. Increased salivation was evident throughout the treatment period, but no alteration of the physical condition was observed. No differences between treated and control animal were detected in body weight development and food consumption. In contrast, water consumption was increased in animals of either sex treated with 1000 mg/Kg bw/day. Increased salivation and increased water consumption might be related to unpalatability problems and irritancy of the stomach.

Microscopic investigations of the stomachs showed epithelial hyperplasia in animals of either sex treated with 1000 mg/Kg bw/day and thickening of the stomach in one female treated with 1000 mg/Kg bw/day at necropsy. The findings are considered a result of local irritation rather than any adverse systemic toxicity of the test item. All parameters related to blood and chemistry examinations and reproductive system showed no treatment-related effects.

 

In view of these results, the NOAEL ‘for systemic toxicity was considered to be 1000 mg/Kg bw/day, and the ‘No Observed Effect Level’ (NOEL) for reproductive toxicity was considered to be 1000 mg/Kg bw/day.

C9 members of the category

 

In a key Guideline (OECD 422) combined repeated dose reproductive/developmental toxicity study (Harlan Laboratories, 2014), the test material (Nonene, branched; CAS# 97280-95-0) was administered by gavage to three groups of Wistar Han™:RccHan™:WIST strain rats (12/sex/dose) for up to eight weeks (including a two week pre-pairing phase, pairing, gestation and early lactation for females), at dose levels of 100, 300 and 1000 mg/Kg bw/day. A control group of twelve males and twelve females was dosed with vehicle alone (Arachis oil BP).

 

Clinical signs, behavioral assessments, body weight change and food and water consumption were monitored during the study. Pairing of animals within each dose group was undertaken on a one male: one female basis within each treatment group on Day 15 of the study, with females subsequently being allowed to litter and rear their offspring to Day 5 of lactation. During the lactation phase, daily clinical observations were performed on all surviving offspring, together with litter size and offspring weights and assessment of surface righting reflex. Extensive functional observations were performed on five selected males from each dose group after the completion of the pairing phase, and for five selected parental females from each dose group on Day 4 post-partum. Hematology and blood chemistry were evaluated prior to termination on five selected males and females from each dose group.

 

Adult males were terminated on Day 43, followed by the termination of all females and offspring on Day 5 post-partum. Any female which did not produce a pregnancy was terminated on or after Day 25 post coitum. One female which did not show positive evidence of mating and did not produce a pregnancy was terminated on Day 57. All animals were subjected to a gross necropsy examination and histopathological evaluation of selected tissues was performed.

 

No test material related mortality was observed through the study period. Clinical signs were detected in animals of either sex treated with 300 and 1000 mg/Kg bw/day during the study. Episodes of increased salivation were reported from Day 8 onwards. The physical condition of males treated with 1000 mg/Kg bw/day was also affected with reductions in body weight development during Weeks 1 and 3 of treatment. Subsequently, a reduction in overall body weight gain in these males and a slight reduction in food efficiency during week 1, was evident. A reduction in haemoglobin was also observed in these males. Water consumption was also significantly increased for animals of either sex from all treatment groups throughout the treatment period. Observations of this nature are often reported when a test material formulation is unpalatable or irritant and can be associated with gastric irritancy rather than be attributable to systemic toxicity. This was supported microscopically with stomach changes identified as epithelial hyperplasia in the forestomach in animals of either sex treated with 1000 mg/Kg bw/day and in males treated with 300 mg/Kg bw/day. This finding was considered to be the result of local irritancy of the test material and therefore cannot be considered indicative of true systemic toxicity.

 

Although there were some statistically significant differences in treated animals from controls for the blood chemical parameters measured, these differences were considered not to be of toxicological significance. Macroscopic findings detected at necropsy were confined to enlarged, pale and mottled kidneys and a dark liver in a number of males treated with 1000 mg/Kg bw/day. One male treated with 300 mg/Kg bw/day also had enlarged kidneys.

 

Histopathological examination of the liver revealed hepatocellular hypertrophy in animals of either sex treated with 1000 and 300 mg/Kg bw/day and in males treated with 100 mg/Kg bw/day. Organ weight data supported this finding with increased absolute and relative liver weights observed in these animals. In the absence of any degenerative or inflammatory changes, this condition is considered to be adaptive in nature.

 

Microscopic examination of the thyroid revealed increased incidence and severity of follicular cell hypertrophy/hyperplasia in animals of either sex treated with 1000 and 300 mg/Kg bw/day and in males treated with 100 mg/Kg bw/day. Males treated with 1000 and 300 mg/Kg bw/day also showed hypertrophic/vacuolated cells in the pituitary. The thyroid, liver and pituitary changes are characteristic of a consequence of hepatocellular induction as a result of enhanced hepatic metabolism. As a side effect of hepatic induction an increased liver metabolism of thyroid hormones T3 and T4 can occur. This subsequently leads to an enhanced thyroid gland production of these hormones as a consequence of a negative feedback stimulation of TSH production. The appearance of thyroid follicular cell hypertrophy and hypertrophic/vacuolated cells in the pituitary are themselves considered to be a result of this process. The thyroid and pituitary changes were considered to be adaptive in nature.

 

Microscopic examination also revealed effects in the kidneys of males from all treatment groups. Tubular basophilia and hyaline droplets was present in males from all treated groups. These tubular findings were also accompanied by tubular degeneration/debris in males treated with 1000 and 300 mg/Kg bw/day. The hyaline droplets can be directly linked to accumulation of alpha 2u-globulin, which is unique to the male rat. This finding is commonly observed in male rats following treatment with some hydrocarbons and is not predictive of any adverse effect in humans. The remaining kidney findings consisting of tubular degeneration/debris may be considered to represent an adverse effect of the test item, however immunohistochemical staining demonstrated that these findings were correlated to the same condition as hyaline droplets.

 

Mating performance and fertility was unaffected by treatment. Offspring viability was however reduced in litters from females treated with 1000 mg/Kg bw/day on Day 4 post-partum. Subsequently reduced litter size and litter weights were evident in these litters on Day 4 post-partum when compared to controls. One female treated with 1000 mg/Kg bw/day also had a total litter loss between Days 2 and 4 post-partum. The mean offspring body weight gains for litters which survived to Day 5 post-partum were also reduced between Days 1 and 4 post-partum at this treatment level.

 

Based on the results observed, the No Observed Adverse Effect Level (NOAEL) for systemic toxicity was determined to be 1000 mg/Kg bw/day. The ‘No Observed Effect Level’ (NOEL) for reproductive toxicity was considered to be 300 mg/Kg bw/day, based on reduced offspring viability, offspring body weight gain, litter size and litter weights on Day 4 post-partum at 1000 mg/Kg bw/day.

C10 members of the category

 

In a supporting Guideline (OECD 422) combined repeated dose, reproductive/developmental toxicity screening test (Harlan Laboratories, 2014), the test material (Decene; CAS# 25339-53-1) was administered by gavage to three groups, each of twelve male and twelve female Wistar Han™:RccHan™:WIST strain rats, for up to eight weeks (including a two week pre-pairing phase, pairing, gestation and early lactation for females), at dose levels of 100, 300 and 1000 mg/Kg bw/day. A control group of twelve males and twelve females was dosed with vehicle alone (Arachis oil BP).

 

Clinical signs, behavioural assessments, body weight change and food and water consumption were monitored during the study.

 

Pairing of animals within each dose group was undertaken on a one male: one female basis within each treatment group on Day 15 of the study, with females subsequently being allowed to litter and rear their offspring to Day 5 of lactation.

 

During the lactation phase, daily clinical observations were performed on all surviving offspring, together with litter size and offspring weights and assessment of surface righting reflex. Extensive functional observations were performed on five selected males from each dose group after the completion of the pairing phase, and for five selected parental females from each dose group on Day 4 post-partum. Haematology and blood chemistry were evaluated prior to termination on five selected males and females from each dose group.

 

Surviving adult males were terminated on Day 43, followed by the termination of all females and surviving offspring on Day 5 post-partum. Any female which did not produce a pregnancy was terminated on or after Day 25 post coitum. Two females showing no visible vaginal opening were terminated on Day 42. All animals were subjected to a gross necropsy examination and histopathological evaluation of selected tissues was performed.

 

No test material related mortality was observed through the study period. Oral gavage administration of the test material at dosages of 100, 300 and 1000 mg/Kg bw/day for approximately six weeks resulted in minimal to moderate microscopic changes in the fore stomach of one male at 300 mg/Kg bw/day and both sexes at 1000 mg/Kg bw/day. These minimal to moderate histopathological changes in the fore stomach were considered to have occurred due to a local irritant effect, rather than true systemic toxicity. As a consequence, the NOAEL for adult systemic toxicity was considered to be 1000 mg/Kg bw/day.

 

There were no treatment-related effects on mating performance; fertility; or gestation length apparent at 100, 300 or 1000 mg/Kg bw/day. There was no effect of treatment on corpora lutea count, pre-implantation loss, numbers of implantations, post-implantation loss, litter size, sex ratio and subsequent offspring survival to Day 4 of age at 100, 300 or 1000 mg/Kg bw/day. Offspring body weight or body weight gain and litter weights, surface righting ability on Day 1 or clinical signs to Day 5 of age remained unaffected post exposure at 100, 300 or 1000 mg/Kg bw/day. Additionally, gross necropsy findings for offspring did not indicate any adverse effect of maternal treatment on offspring development at 100, 300 or 1000 mg/Kg bw/day.

 

Based on the results observed, the No Observed Effect Level (NOEL) for reproduction, including the survival, growth and development of the offspring, is considered to be 1000 mg/Kg bw/day.

 

C14 members of the category

In a supporting Guideline (OECD 422) combined repeated dose, reproductive/developmental toxicity study (Daniel, 1995), the test material (1-Tetradecene; CAS# 1120-36-1) was administered via oral gavage to Sprague-Dawley Crl:CD®BR VAF/Plus®rats (12 males and 20 females/dose) at doses of 0, 100, 500, or 1000 mg/Kg/day in corn oil. The F0 females were treated for 28 days prior to mating until the day prior to euthanasia (43-47 days). A satellite group of F0 females were added for neurotoxicity, clinical pathology and histopathological evaluations. Twelve F0 females assigned to a breeding phase were treated for 14 days prior to mating, during mating, gestation, and lactation until the day prior to euthanasia (42-51 days).

 

Animals were observed daily for overt signs of toxicity, and body weights and food consumption were measured at specific intervals. Breeding females were allowed to deliver and rear their offspring until lactation Day 4. Viability and development of the F1 generation were evaluated and surviving F1 pups were euthanized and necropsied on lactation Day 4. Functional observation battery, motor activity and clinical pathology evaluations were performed on selected F0 males and all satellite females. All F0 males and females were subjected to gross necropsy at the time of euthanasia. Specified tissues were retained at necropsy for selected F0 males and all (satellite and breeding) females and preserved in Bouin’s solution or 10% neutral buffered formalin. All tissues and organs from five randomly selected males and females from the control and high dose groups; lungs, liver, kidneys, and reproductive tract from all females; and gross lesions from all animals in each group were processed for microscopic examination.

 

Oral administration of the test material to virgin females and F0 parental animals produced minor clinical signs (salivation and urine staining) and dose-related hydrocarbon nephropathy in male rats in the 100, 500, and 1000 mg/Kg/day groups. This finding in male rats is commonly associated with hydrocarbon nephropathy and is not believed to pose any risk to humans.

 

In the satellite females, minor decreases were observed in the mean red cell count and hematocrit in 100, 500 and 1000 mg/Kg/day groups, as well as in haemoglobin and MCV in the 1000 mg/Kg/day group. Similar changes, though no statistically significant were observed in males. Therefore, the biological significance of these changes in the females is not clear.

 

In males, statistically significant minor increases in ALT were observed in 500 and 1000 mg/Kg/day groups. In satellite females, there were statistically significant decreases in sodium in 100, 500 and 1000 mg/Kg/day groups and increases in cholesterol in the 500 and 1000 mg/Kg/day groups. The changes of ALT sodium and cholesterol were observed in one sex and therefore were considered of no biological significance.

 

Minimal to moderate hepatocyte cytoplasmic vacuolation was observed in the livers of male and female rats at dosages of 500 and 1000 mg/Kg/day and correlated with increased liver weights at same tested doses. However, only the statistically significant increase of liver weight to brain weight ratio was seen in 1000 mg/Kg/day group females. The hepatocellular vacuolation was clearly related to the test item administration. However, in the absence of any other histopathological changes in liver as well as in the absence of significant effects on liver function (as indicated by the measured clinical chemistry parameters) the hepatocellular vacuolation might be considered to represent an adaptive response.

 

There were no other apparent test-article related histopathologic findings seen in any other tissues or organs examined. A statistically significant decrease in spleen weight relative to brain weight was noted in the 1000 mg/Kg/day group females. In addition, a statistically significant increase in absolute kidney weight was observed in 500 mg/Kg/day group females. These changes in female organs were not believed to be biologically meaningful because no test article-related lesions were seen histologically in these organs.

 

There was no evidence of impaired reproductive capabilities in the F0 generation or developmental toxicity in the F1 generation through lactation Day 4. The pup survival, litter sizes, sex ratios, external observations, body weights and necropsy of the F1 pups were comparable between all the groups.

 

Based on the results observed, the NOAEL for systemic toxicity was determined to be 100 mg/Kg/day in satellite females. Since hydrocarbon nephropathy was observed at all dose levels in male rats, a NOAEL for systemic toxicity could not be determined for males. The NOAEL for reproductive, developmental, or neurotoxicity was determined to be 1000 mg/Kg/day in male and female rats.

C16 members of the category

 

In a supporting Guideline (OECD 422) combined repeated dose, reproductive/developmental toxicity study (Harlan Laboratories, 2014), the test material (Hexadecene; CAS# 26952-14-7) was administered by gavage to three groups, each of twelve male and twelve female Wistar Han™:RccHan™:WIST strain rats, for approximately six weeks (including a two week pre-pairing phase, pairing, gestation and early lactation for females), at dose levels of 100, 300 and 1000 mg/Kg bw/day. A control group of twelve males and twelve females was dosed with vehicle alone (Arachis oil BP).

 

Clinical signs, behavioral assessments, body weight change and food and water consumption were monitored during the study. Pairing of animals within each dose group was undertaken on a one male: one female basis within each treatment group on Day 15 of the study, with females subsequently being allowed to litter and rear their offspring to Day 5 of lactation.

 

During the lactation phase, daily clinical observations were performed on all surviving offspring, together with litter size and offspring weights and assessment of surface righting reflex. Extensive functional observations were performed on five selected males from each dose group after the completion of the pairing phase, and for five selected parental females from each dose group on Day 4 post-partum. Hematology and blood chemistry were evaluated prior to termination on five selected males and females from each dose group.

 

Adult males were terminated on Day 43, followed by the termination of all females and offspring on Day 5 post-partum. Any female which did not produce a pregnancy was terminated on or after Day 25 post coitum. One female which did not show positive evidence of mating and did not produce a pregnancy was terminated on Day 57. All animals were subjected to a gross necropsy examination and histopathological evaluation of selected tissues was performed. No mortality was observed through the study period. Oral gavage administration of the test material at dosages up to 1000 mg/Kg bw/day for approximately six weeks (including two weeks pre-pairing, gestation and early lactation for females) was well tolerated, with no effect of treatment for body weight performance or food intake being apparent in either sex.

 

Treatment related clinical signs were restricted to increased post dosing salivation for both sexes at 1000 mg/Kg bw/day, and a few females at 300 mg/Kg bw/day but this was considered to reflect palatability of the test item rather than any systemic effect of treatment. Increased water consumption, compared to control, was apparent for males at 1000 mg/Kg bw/day, which may also reflect palatability of the test item. Assessment of hematology and blood chemistry parameters did not reveal any differences considered to indicate an effect of treatment at dosages up to 1000 mg/Kg bw/day. Macroscopic necropsy findings and organ weights also did not indicate any treatment related effects. Histopathological examination of tissues on the study did not reveal any findings that were considered to indicate systemic toxicity. At 1000 mg/Kg bw/day, minimal or moderate peribronchiolar inflammation was observed in the lungs for both sexes, but this was considered to be a consequence of accidental aspiration of the test item during the gavage procedure and to be of no toxicological significance. There were no effects of treatment on the reproductive performance, including offspring survival, growth and development, at 100, 300 or 1000 mg/Kg bw/day.

 

Based on the results observed, 1000 mg/Kg bw/day was considered to represent the No Observed Effect Level (NOEL) for adult toxicity as well as a clear No Observed Adverse Effect Level (NOAEL).The No Observed Effect Level (NOEL) for reproduction, including the survival, growth and development of the offspring, is also considered to be 1000 mg/Kg bw/day.

 

C18 members of the category

In a supporting Guideline (OECD 422) repeated dose, reproductive/developmental toxicity screening study (Harlan Laboratories, 2014), the test material (Octadec-1-ene UVCB; CAS# 112-88-9) was administered by gavage to Wistar Han™: RccHan™: WISTstrain rats (12/sex/dose), for up to eight weeks (including a two-week pre-pairing phase, pairing, gestation and early lactation for females), at dose levels of 100, 300 and 1000 mg/Kg bw/day. A control group of twelve males and twelve females was dosed with vehicle alone (Arachis oil BP).

 

Clinical signs, behavioral assessments, body weight change and food and water consumption were monitored during the study. Pairing of animals within each dose group was undertaken on a one male: one female basis within each treatment group on Day 15 of the study, with females subsequently being allowed to litter and rear their offspring to Day 5 of lactation. During the lactation phase, daily clinical observations were performed on all surviving offspring, together with litter size and offspring weights and assessment of surface righting reflex. Extensive functional observations were performed on five selected males from each dose group after the completion of the pairing phase, and for five selected parental females from each dose group on Day 4 post-partum. Hematology and blood chemistry were evaluated prior to termination on five selected males and females from each dose group. Adult males were terminated on Day 43, followed by the termination of all females and offspring on Day 5 post-partum. Any female which did not produce a pregnancy was terminated on or after Day 25 post coitum. All animals were subjected to a gross necropsy examination and histopathological evaluation of selected tissues was performed.

 

No mortality was observed through the study period. Clinical signs were confined to increased salivation detected in animals of either sex treated with 1000 and 300 mg/Kg bw/day. Observations of this nature are commonly observed following the oral administration of an unpalatable or slightly irritant test item formulation and do not necessarily represent an adverse effect of treatment.

 

No adverse effect was apparent for body weight or food consumption in treated animals throughout the treatment period. Although there were some statistically significant differences in treated animals from controls for the hematological and blood chemical parameters measured, these differences were considered not to be of toxicological significance. No macroscopic abnormalities were evident in treated animals; however microscopic examination revealed changes in the mesenteric lymph nodes and spleen. A minimal or mild inflammatory cell infiltrate was present in the periglandular fat surrounding the mesenteric lymph nodes in animals of either sex treated with 1000 and 300 mg/Kg bw/day. The infiltrate was mixed in character, principally composed of lymphocytes and neutrophils and tended to have a perivascular or perilymphatic orientation. Although the origin of the finding is uncertain it might have resulted from local irritation of material from the gut reaching the drainage nodes however there was no evidence of local irritation to the gut mucosa. However, in the absence of any other associated pathological changes at the low severity levels noted the change is considered likely to be non-adverse. The statement of laboratory is provided as an attachment to this IUCLID record.

The incidence and severity of extramedullary hematopoiesis was increased in the spleen in animals of either sex treated with 1000 mg/Kg bw/day and in females treated with 300 mg/Kg bw/day. The inter-group differences in females were not clearly dosage-related and taking into account the low number of animals examined, the absence of supportive intergroup differences in hematological values or bone marrow findings, these differences were considered more likely to have arisen as a result of individual variation, or possibly as a response to blood loss during parturition, than to be test item related. There were no statistically significant differences in adult organ weights that, in the absence of any evidence of histopathological change, were considered to be of any toxicological significance.

 

Mating performance and fertility was unaffected by treatment and there were no treatment-related effects detected in the offspring parameters observed.

 

Based on the results observed, the ‘No Observed Effect Level’ (NOEL) for systemic toxicity was considered to be 100 mg/Kg bw/day and the ‘No NOEL for reproductive toxicity was considered to be 1000 mg/Kg bw/day. 

 

In a reproduction/developmental screening study (Thorsud, 2003), octadecenes, dissolved in corn oil, was administered to Sprague-Dawley rats (12 sex/dose) by gavage at dose levels of 0, 100, 500, or 1000 mg/Kg/day for 42 days.

 

Parents and offspring were observed for clinical signs of toxicity, mortality, mating, parturition, lactation, off-spring growth and maturity. Body weight and food consumption determinations were also conducted periodically through the study period. All parents and offspring were subjected to necroscopy following termination or death. 

 

There was no mortality observed in animals in the control, 100, 500, or 1000 mg/Kg dose group. No treatment-related or dose-dependant signs of clinical toxicity were noted in rats at any dose level. Mean body weight, body weight change and food consumption was observed to be normal in all treatment animals when compared with the controls. Parent female mating, fertility, and mean gestation lengths were observed to be comparable with controls as were the mean number of pups delivered and live birth and viability indices. Mean live pups/litter and sex ratio/litter were also found to be normal and comparable to those observed in control animals.

 

Gross necroscopy revealed no remarkable differences between octadecenes-treated and control animals. There were no microscopic lesions observed in male or female rats treated with octadecenes and no statistically significant differences in absolute or relative epididymides weight were noted in treated males when compared with control males. There were no statistically significant differences observed in pup weights on lactation days 1 and 4 and gross necroscopy on lactation day 4 revealed no treatment-related effects.

 

Based on the lack of adverse systemic effects observed in the study, the developmental/reproductive toxicity NOAEL for octadecene was reported as 1000 mg/Kg/day.

Additionally, OECD 443 test proposals for the substances hex-1-ene, hexadecene, octadec-1-ene, alkenes C8-10, C9-rich, and nonene, branched have been proposed based on structural characteristics and the results of data available for higher olefin category members on uptake following oral administration, together with repeated dose and developmental toxicity. It is proposed that the studies will be carried out in rats using oral (gavage) administration and will follow the basic study design outlined in EU method B.56/OECD 443.

 

Supporting Inhalation Toxicity Data

Supporting information is also available from a sub-chronic repeat dose inhalation toxicity study (Bennick et al., 1984) where 1-hexene was administered to Fischer 344 rats (40/sex/concentration) by whole body exposure at concentrations of 0, 300, 1000, or 3000 parts per million (corresponding to 0; 1033; 3442; or 10,326 mg/m3), 6 hours a day, 5 days a week, for a period of 13 weeks. Ten rats/ sex/ concentration were used for neuromuscular testing, ten rats/ sex/ concentration were sacrificed after 7 weeks of exposure, and 20 rats/ sex/ concentration were sacrificed after 13 weeks of exposure.

 

Sub-chronic inhalation of 1-hexene for 13 weeks did not produce any adverse reproductive or testicular effects in rats. The NOAEC was determined to be 3000 ppm (10,326 mg/m3) based on the lack of toxicologically relevant findings at the highest concentration tested.

Justification for selection of Effect on fertility via oral route:
Oral screening level information, supported by results from a 90-day sub-chronic inhalation study, is available on the potential of 4 members of this category covering C6 to C18 to affect reproduction and fertility. No effects were observed in male or female rats receiving oral treatments at dose levels up to 1000 mg/kg bw/day, or inhalation exposures up to 10,326 mg/m3.

Effects on developmental toxicity

Description of key information

Guideline (OECD 414) developmental toxicity studies have been conducted for five members of the higher olefins category.

No effects of the development of the off-spring were observed in any study at dose levels up to 1000 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:
11 February 2015 - 26 March 2015
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: Study conducted according to OECD 414 Guideline and GLP conditions. Fully adequate for assessment.
Justification for type of information:
A discussion and report on the read across strategy is given as an attachment in Section 13.
Reason / purpose:
read-across: supporting information
Qualifier:
according to
Guideline:
OECD Guideline 414 (Prenatal Developmental Toxicity Study)
Deviations:
yes
Remarks:
Deviation did not affect the purpose or integrity of the study.
Qualifier:
according to
Guideline:
EPA OPPTS 870.3700 (Prenatal Developmental Toxicity Study)
Qualifier:
according to
Guideline:
other: Japanese Ministry of Agriculture, Forestry and Fisheries Testing guidelines for Toxicology studies, 12 NohSan No 8147, (24 November 2000)
Qualifier:
according to
Guideline:
other: Commission Regulation (EC) No 440/2008 of 30 May 2008 test methods pursuant to Regulation (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)
Specific details on test material used for the study:
- Test Material: Octadecene
- CAS Number: CAS 27070-58-2
- Physical State/Appearance : Clear colorless liquid
- Molecular Formula: C18 Isomerized Olefin
- Purity: 94.1%
- Batch Number: 747273
- Label: C18 Isomerised Olefin Lot 747273
- Date Received: 3 March 2014
- Storage Conditions: Room temperature, in the dark and under nitrogen (after opening the container)
- Expiry Date: 10 February 2016

Octadecene used in this study was a typical production sample, according to the details included in the boundary composition in IUCLID section 1.2. Octadecene was chosen in the Higher Olefins category testing strategy because it represents a substance with high di-sub content (category range 0.3 – 94%) and high tri-sub content (category range 1 - 65%). Please see the testing strategy attached in section 13 for further details.
Species:
rat
Strain:
other: Sprague-Dawley Crl:CD (SD) IGS BR strain
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Charles River (UK) Limited, Margate, Kent
- Age at study initiation: not specified
- Weight at study initiation: 180 to 284g.
- Housing: solid-floor polypropylene cages with stainless steel mesh lids furnished with softwood flakes
- Diet (e.g. ad libitum): A pelleted diet (Rodent 2018C Teklad Global Certified Diet, Harlan UK, Oxon, UK) - free access
- Water (e.g. ad libitum): freely available


ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22 ± 3 ºC
- Humidity (%): 50 ± 20%
- Air changes (per hr): 15 air changes per hr
- Photoperiod (hrs dark / hrs light): 12 hr light/12 hr dark

Experimental Starting Date: 11 February 2015
Experimental Completion Date: 26 March 2015

Justification for specie selection: the selected species is a readily available rodent species historically used in safety evaluation studies and is acceptable to appropriate regulatory authorities.


IN-LIFE DATES: From 11 February 2015 to 26 March 2015
Route of administration:
oral: gavage
Vehicle:
arachis oil
Details on exposure:
DIET PREPARATION
- Rate of preparation of diet (frequency): Formulations were prepared once.
- Mixing appropriate amounts with (Type of food): The appropriate concentrations were prepared in Arachis oil solutions.
- Storage temperature of food: Stored at approximately +4 °C in the dark.

Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
The stability and homogeneity of the test item formulations were previously determined by Harlan Laboratories Ltd., Shardlow, UK Analytical Services (Project Number 41301656 - Octadecene CAS 27070-58-2: Ninety Day Repeated Dose Oral (Gavage) Toxicity Study in Rats). The results indicate that the prepared formulations were within 90-103% of the nominal concentration confirming the accuracy of the formulation procedure.
Details on mating procedure:
Female animals were delivered in two batches 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:
From Day 5 to Day 19 of gestation, by gavage
Frequency of treatment:
Daily
Duration of test:
From Day 5 to Day 19 of gestation
Dose / conc.:
100 mg/kg bw/day
Dose / conc.:
300 mg/kg bw/day
Dose / conc.:
1 000 mg/kg bw/day
No. of animals per sex per dose:
24 Females
Control animals:
yes, concurrent vehicle
Details on study design:
- Dose selection rationale: Dose levels were selected based on available toxicity data including a dose range finding study (Harlan Laboratories Ltd report number 41403655)
Maternal examinations:
CAGE SIDE OBSERVATIONS:
Once daily during the gestation period. During the dosing period, observations were recorded immediately before and soon after dosing and one hour post dosing.

BODY WEIGHT:
Individual body weights were recorded on Day 3 (prior to dosing) and on Days 5, 6, 7, 8, 11, 14 and 17 of gestation of gestation, including for surviving animals at terminal kill (Day 20).

FOOD CONSUMPTION AND COMPOUND INTAKE (if feeding study):
Food consumption was recorded for each surviving individual animal at Day 3, 5, 8, 11, 14, 17 and 20 of gestation.

WATER CONSUMPTION AND COMPOUND INTAKE (if drinking water study):
Daily by visual inspection of the water bottles for any overt changes.

POST-MORTEM EXAMINATIONS: Yes
- Full external and internal examination ( any macroscopic abnormalities were recorded).

Ovaries and uterine content:
Ovaries and uteri examinations:
i) Number of corpora lutea
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
Fetal examinations:
- Fetal external findings
- Visceral findings
- Skeletal findings and skeletal development

- Number of implantations
- Embryofetal survival
- Litter size
- Sex ratio
- Mean fetal litter and placental weights


Statistics:
The following parameters were analyzed statistically, where appropriate, using the test methods outlined below:
Body weight and body weight change (including adjustment for the contribution of the gravid uterus), food consumption, gravid uterus weight, litter data and fetal litter and placental weights.
Data were first analysed using Shapiro Wilk normality test and Bartlett’s test for homogeneity of variance. Where there was no significance, parametric methodology was applied using one way analysis of variance and, if significant, Dunnett’s multiple comparison test. Where statistical significance was observed, non parametric methodology was applied using Kruskal-Wallis nonparametric analysis of variance; and, if significant, pairwise analysis of control values against treated values using the Mann-Whitney ‘U’ test. Due to the preponderance of non-normal distributions for litter/fetal parameters, these data were routinely analyzed using non-parametric methodology.

Fetal morphology parameters, including skeletal or visceral findings were generally analysed by Kruskal- Wallis and, if significant, Mann-Whitney ‘U’ test.

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
Percentage post-implantation loss
Sex ratio
Clinical signs:
effects observed, non-treatment-related
Description (incidence and severity):
No clinical signs of toxicity were detected.

One control female and one female treated with 300 mg/kg bw/day exhibited generalized fur loss. A further animal treated with 300 mg/kg bw/day also exhibited fur staining. In the absence of findings for high dose females, these observations were considered to be incidental and unrelated to treatment.
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, were unaffected by treatment at 100, 300 or 1000 mg/kg bw/day.
Food consumption and compound intake (if feeding study):
no effects observed
Description (incidence and severity):
Food consumption during gestation was unaffected by treatment at 100, 300 or 1000 mg/kg bw/day.
Food efficiency:
not specified
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, non-treatment-related
Description (incidence and severity):
Findings observed at macroscopic necropsy did not indicate any effect of treatment. One control female exhibited generalized fur loss. In the absence of similar findings in high dose females this observation was considered to be incidental and unrelated to treatment.
Neuropathological findings:
not examined
Histopathological findings: non-neoplastic:
not examined
Histopathological findings: neoplastic:
not examined
Other effects:
not examined
Other effects:
no effects observed
Description (incidence and severity):
The number of implantations, subsequent embryofetal survival and litter size, sex ratio and mean fetal, litter and placental weights on Day 20 of gestation were unaffected by maternal treatment at 100, 300 and 1000 mg/kg bw/day.
Details on maternal toxic effects:
Maternal toxic effects:no effects

Details on maternal toxic effects:
No maternal toxicity
Key result
Dose descriptor:
NOEL
Effect level:
1 000 mg/kg bw/day (nominal)
Based on:
test mat.
Basis for effect level:
other: Systemic Toxicity
Key result
Abnormalities:
no effects observed
Fetal body weight changes:
no effects observed
Description (incidence and severity):
Mean fetal, litter and placental weights on Day 20 of gestation were unaffected by maternal treatment at 100, 300 and 1000 mg/kg bw/day.
Migrated Data from removed field(s)
Field "Fetal/pup body weight changes" (Path: ENDPOINT_STUDY_RECORD.DevelopmentalToxicityTeratogenicity.ResultsAndDiscussion.ResultsFetuses.FetalPupBodyWeightChanges): no effects observed
Field "Description (incidence and severity)" (Path: ENDPOINT_STUDY_RECORD.DevelopmentalToxicityTeratogenicity.ResultsAndDiscussion.ResultsFetuses.DescriptionIncidenceAndSeverityFetalPupBodyWeightChanges): Mean fetal, litter and placental weights on Day 20 of gestation were unaffected by maternal treatment at 100, 300 and 1000 mg/kg bw/day.
Reduction in number of live offspring:
no effects observed
Changes in sex ratio:
no effects observed
Description (incidence and severity):
Sex ratio on Day 20 of gestation was unaffected by maternal treatment at 100, 300 and 1000 mg/kg bw/day.
Changes in litter size and weights:
no effects observed
Description (incidence and severity):
Litter size and weights on Day 20 of gestation were unaffected by maternal treatment at 100, 300 and 1000 mg/kg bw/day.
Changes in postnatal survival:
no effects observed
Description (incidence and severity):
Embryofetal survival was unaffected by maternal treatment at 100, 300 and 1000 mg/kg bw/day.
External malformations:
no effects observed
Description (incidence and severity):
Neither the type, incidence or distribution of findings observed during external examination of the fetuses at necropsy on Day 20 of gestation indicated any adverse effect of maternal treatment on fetal development.
Skeletal malformations:
effects observed, treatment-related
Description (incidence and severity):
Neither the type, incidence or distribution of findings observed during external examination of the fetuses at necropsy on Day 20 of gestation and subsequent detailed visceral and skeletal examination indicated any adverse effect of maternal treatment on fetal development.

Intergroup differences for some skeletal parametersoccasionally attained statistical significance but these isolated differences were considered incidental and did not indicate any disturbance of fetal development.
Visceral malformations:
no effects observed
Description (incidence and severity):
Neither the type, incidence or distribution of findings observed during external examination of the fetuses at necropsy on Day 20 of gestation and subsequent detailed visceral and skeletal examination indicated any adverse effect of maternal treatment on fetal development.
Other effects:
no effects observed
Details on embryotoxic / teratogenic effects:
Embryotoxic / teratogenic effects:no effects

Details on embryotoxic / teratogenic effects:
No embryotoxic/teratogenic effects
Key result
Dose descriptor:
NOEL
Effect level:
1 000 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 2. Summary of Female Performance

Category

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

0 (control)

100

300

1000

Initial Group Size

24

24

24

24

Pregnant

24

24

24

24

 

Table 3. 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

Body Weight

Change (g)

during Days of

Gestation

Gravid

Uterus

Weight

(g)

Adjusted

Body

Weight (g)

Day 20

Adjusted

Body Weight

(g)

Change

 

5

20

5-20

Day 20

5-20

0

(control)

Mean

255.7

374.5

118.8

83.667

290.8

35.1

Sd

30.7

48.6

21.4

12.753

39.0

12.5

n

24

24

24

24

24

24

 

100

Mean

256.7

381.8

125.0

86.848

294.9

38.2

Sd

22.4

31.2

18.5

11.060

26.7

13.2

n

24

24

24

24

24

24

 

300

Mean

258.8

380.0

121.2

87.820

292.2

33.4

Sd

20.4

24.7

14.1

8.848

22.8

10.6

n

24

24

24

24

24

24

 

1000

Mean

257.8

374.1

116.3

81.928

292.2

34.3

Sd

24.6

39.6

19.9

15.792

31.6

13.0

n

24

24

24

24

24

24

Table 4. 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

13.9

13.5

0.2

0.2

0.4

2.3

2.9

7.3

5.8

13.2

55.3

4.185

3.981

4.092

0.569

53.555

7.478

Sd

2.0

1.9

0.5

0.7

1.1

3.9

8.1

2.0

1.5

2.3

9.7

0.255

0.273

0.253

0.057

7.966

1.392

n

24

24

24

24

24

24

24

24

24

24

24

24

24

24

24

24

24

 

100

Mean

14.3

14.0

0.2

0.0

0.2

2.4

1.6

6.5

7.3

13.8

47.2

4.122

3.917

4.012

0.559

55.012

7.656

Sd

1.9

1.9

0.5

0.2

0.7

5.4

5.0

2.2

2.1

2.0

13.7

0.218

0.196

0.196

0.048

7.555

1.166

n

24

24

24

24

24

24

24

24

24

24

24

24

24

24

24

24

24

 

300

Mean

14.7

14.3

0.1

0.0

0.1

2.2

2.1

7.5

6.5

14.0

53.4

4.126

3.909

4.027

0.563

56.340

7.873

Sd

1.4

1.5

0.4

0.0

0.4

4.3

6.7

1.9

2.0

1.5

13.6

0.194

0.181

0.177

0.061

6.343

1.098

n

24

24

24

24

24

24

24

24

24

24

24

24

24

24

24

24

24

 

1000

Mean

13.9

13.5

0.4

0.0

0.5

3.4

3.6

6.3

6.7

13.0

49.0

4.127

3.925

4.023

0.557

52.280

7.200

Sd

1.7

2.4

0.8

0.2

0.8

11.2

6.6

1.9

2.1

2.6

11.4

0.250

0.214

0.232

0.059

10.888

1.550

n

24

24

24

24

24

24

24

24

24

24

24

24

24

24

24

24

24

Table 5. Summary Incidence of Fetal Skeletal Findings

Skeletal

Findings

Dose Level (mg/kg bw/day)

0 (control)

100

300

1000

Number of Fetuses (litters) Examined

150 (24)

160 (24)

162 (24)

150 (24)

NF

NL

%†

NF

NL

%†

NF

NL

%†

NF

NL

%†

Skull

 

Hyoid - incomplete ossification

21

13

14.0

22

13

13.8

12

8

7.2

6

6

3.8*

Vertebral Column

 

Number of pre-sacral vertebrae = 25/27

0

0

0.0

0

0

0.0

4

4

2.7*

0

0

0.0

* Significantly different from control group p<0.05

Table 6. Historical Data - Charles River (UK) Limited - Normal Ranges for Pre-Natal Study Skeletal Fetal Findings in the Sprague-Dawley Crl:CD®(SD) IGS BR Rat

Skeletal findings

Range **

Group mean % of fetuses affected

No. of Litters

{fetuses} examined

Hyoid - incomplete ossification

5 (11.5) - 18.4 [4.0]

255 {1605}

 

Number of pre-sacral vertebrae = 25/27

0 (0.4) – 2.7 [0.8]

255 {1605}

** Range = minimum to maximum value ( ) = group mean  [ ] = 1 standard deviation

 

Where the range exceeds the minimum or maximum possible physiological values, the minimum or maximum physiological values are presented.

Data shown are recorded for presentation purposes, therefore it is not always possible to calculate the range exactly from the mean and SD presented.


Conclusions:
Based on the result of the study, the ‘No Observed Effect Level' (NOEL) for the pregnant female and the ‘No Observed Effect Level' (NOEL) for developmental toxicity is considered to be 1000 mg/kg bw/day.
Executive summary:

In a key Guideline (OECD 414) pre-natal developmental toxicity study, the test material (Octadecene; CAS# 27070-58-2) was administered by gavage to three groups each of twenty-four time mated Sprague-Dawley Crl:CD® (SD) IGS BR strain rats, between Days 5 and 19 of gestation inclusive at dose levels 100, 300, and 1000 mg/Kg bw/day. A further group of twenty-four time mated females was exposed to the vehicle only (Arachis Oil BP) to serve as a control over the same treatment period.

 

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.

 

No mortality was observed through the study period and oral administration of the test material to pregnant rats by oral gavage during gestation at dose levels of 100, 300 and 1000 mg/Kg bw/day did not result in any treatment related effects. The No

Observed Effect Level' (NOEL) for the pregnant female was considered to be 1000 mg/Kg bw/day.

 

No treatment-related changes were detected in the offspring parameters measured. The ‘No Observed Effect Level’ (NOEL) for developmental toxicity was therefore considered to be 1000 mg/Kg bw/day.

Effect on developmental toxicity: via oral route
Endpoint conclusion:
no adverse effect observed
Dose descriptor:
NOAEL
1 000 mg/kg bw/day
Study duration:
subchronic
Species:
rat
Quality of whole database:
In total, fourteen good quality guideline and screening studies are available for ten category members, covering C6 to C30 higher olefins. The study on Octadecene is one of five good quality, guideline reproductive toxicity studies conducted on members of the higher olefins category. In addition, nine supporting toxicity screening studies are available for category members. None of the studies showed any evidence of effects on development of the rat foetus at levels up to 1000 mg/Kg.
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

Developmental Toxicity

 

Guideline (OECD 414, 421 and 422) developmental toxicity studies are available for the following members of the higher olefin Category:

Test substance identity

Testing for Developmental Effects

Hex-1-ene

OECD 414 & OECD 421

Alkenes, C6

OECD 422

Oct-1-ene

OECD 422

Nonene

Branched

OECD 414 & OECD 422

Decene

OECD 422

Tetradec-1-ene

OECD 422

Hexadecene

OECD 422

Octadec-1-ene

OECD 414 & OECD 422

Octadecene

OECD 414 & OECD 421

Hydrocarbons, C12-30 Olefin rich

OECD 414

Information is available from multiple guideline (OECD 414, 421 AND 422) studies that investigated the developmental toxicity potential in rats, for members of the higher olefin category. Studies covered substances in the range C6 to C12-30, following oral exposure.

C6 members of the category

In a supporting Guideline (OECD 414) pre-natal developmental toxicity study (Envigo Research Ltd., 2016), the test material (hex-1-ene; CAS# 592-41-6) 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 100, 300, and 1000 mg/Kg bw/day. A further group of twenty-four time mated females was exposed to the vehicle only (Arachis oil BP) 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.

 

No mortality was observed through the study period and oral administration of the test material to pregnant rats by oral gavage during gestation at dose levels of 100, 300 and 1000 mg/kg bw/day did not result in any treatment related effects.

 

The ‘No Observed Effect Level’ (NOEL) for the pregnant female was considered to be 1000 mg/Kg bw/day. No treatment-related changes were detected in the offspring parameters measured. The ‘No Observed Effect Level’ (NOEL) for developmental toxicity was therefore considered to be 1000 mg/Kg bw/day.

 

In a supporting Guideline (OECD 422) combined repeated dose, reproductive/developmental toxicity screening study , (Thorsud, 2003), the test material (C6-Olefin/Hexene) was administered to Sprague Dawley CrI:CD®(SD)IGS BR rats (12/sex/dose) once daily (seven days a week) by oral gavage at doses of 0, 100, 500, or 1000 mg/Kg/day for a minimum of 14 days prior to mating and continuing through lactation day 3.

 

Cage-side observations for overt signs of toxicity were performed once daily within approximately one-half hour to two hours following dosing. Detailed clinical observations were performed a minimum of weekly until evidence of mating was detected, daily during gestation and lactation and on the day of scheduled euthanasia. Individual body weights were recorded on days 0, 3, 7 and 12 prior to mating, on gestation days 0, 7, 14 and 20 and on lactation days 1 and 4. Individual food consumption was recorded on the same days as body weights (except during cohabitation). After a minimum of 14 days of treatment, females were cohabited with males from the same treatment group in the toxicity study.

Following parturition, pup viability, sex determinations, observations and body weights were recorded at specified intervals during lactation days 0 and 4. Surviving females and their pups were euthanized and necropsied on lactation day 4. Fresh organ weights were recorded for surviving females and selected tissues and organs were preserved from all females. All gross lesions collected at necropsy from all females were examined microscopically.

 

There were no toxicologically meaningful differences noted in F0 mean body weights, body weight gain or food consumption between the control and test article-treated groups. The F0 female mating and fertility indices and mean gestation lengths were comparable between the control and test article-treated groups. The mean number of F1 pups delivered and the F1 live birth index were comparable between the control and test article-treated groups, and there were no toxicologically meaningful differences in the F1 viability index (pups surviving to lactation day 4) between the groups. The mean F1 pups per litter and the F1 pup sex ratio were comparable between the control and test article-treated groups on lactation days 0 and 4.

 

Mean F1 pup weights were slightly but not statistically lower than controls in the 500 mg/Kg/day group on lactation day 1 and in the 1000 mg/Kg/day group on lactation days 1 and 4. However, the mean body weights in these groups were within the range of SLI historical control data. Mean F1 pup weights in the 100 mg/Kg/day group were comparable to controls on lactation days 1 and 4. There were no toxicologically meaningful differences in pup observations during lactation.

 

No other remarkable findings were noted in the F1 pups during lactation. No remarkable gross necropsy findings were noted for F1 pups found dead during the study or euthanized at study termination. No remarkable internal gross necropsy findings were noted for surviving F0 females in the control or test article-treated groups at study termination. There were no toxicologically meaningful differences in F0 absolute or relative organ weight data between the groups and no toxicologically meaningful microscopic findings were noted for F0 females in the test article-treated groups.

 

Based on the results of this study, a dosage level of 1000 mg/Kg/day was considered a no-observed-adverse-effect level (NOAEL) for developmental toxicity. 

 

Additionally, in an OECD 421 screening for reproductive/developmental toxicity study (Daniel, 1995), 1-hexene was administered via gavage to twelve Sprague-Dawley rats/sex/dose at doses of 0, 100, 500, or 1000 mg/Kg/day. The test material was composed of equal amounts of Neodene 6 alpha olefin, hexene-1 Gulftene 6, and alpha olefin C6 1 -hexene, which were obtained from different sources. Males were treated for 44 days beginning 28 days prior to mating and females were treated for 41 to 55 days beginning 14 days prior to mating through lactation day 4.

 

No reproductive or developmental effects were observed. There was a slight, but significant, decrease in absolute epididymal weight at all concentrations. The relative epididymal to brain weight was only significantly decreased in the low-dose group. Although the absolute epididymides weight was significantly decreased in parental males; the change was within 10% of the control, there was no dose response, there was no effects noted microscopically, and there were no effects on fertility. Therefore, this is not considered to be toxicologically significant. Pitted kidneys were observed at necropsy for 2 of 12 mid-dose males and 3 of 12 high-dose males. The predominant microscopic finding in males was the presence of large hyaline droplets in the proximal convoluted tubule that was dose related. These findings suggest hydrocarbon nephropathy, which is a toxicological effect specific to male rats and is not considered relevant to humans. There was no LOAEL for this study.

 

The NOAEL for systemic, reproductive, and developmental toxicity was 1000 mg/Kg/day, which excluded the hydrocarbon nephropathy in males.

C8 member of the category

 

An Oral (Gavage) Combined Repeat Dose Toxicity Study with Reproduction/Developmental Toxicity Screening Test in the Rat (OECD 422) was performed on the test material Oct-1-ene CAS 111-66-0 (Harlan Laboratories Ltd., 2014).

 

After eight weeks treatment clinical signs were detected in animals of either sex treated with 1000 mg/Kg bw/day and in males treated with 300 mg/Kg bw/day. Increased salivation was evident throughout the treatment period, but no alteration of the physical condition was observed. No difference between animals treated and controls were detected in body weight development and food consumption. In contrast, the water consumption was increased in animals of either sex treated with 1000 mg/Kg bw/day. Increased salivation and increased water consumption might be related to unpalatability problems and irritancy of the stomach. Microscopic investigations of the stomachs showed epithelial hyperplasia in animals of either sex treated with 1000 mg/Kg bw/day and thickening of the stomach in one female treated with 1000 mg/Kg bw/day at necropsy. The findings are considered a result of local irritation rather than any adverse systemic toxicity of the test item. All parameters related to blood and chemistry examinations and reproductive system showed no treatment-related effects.

No significant differences were detected for corpora lutea, implantation counts, implantation losses, litter size or litter viability for treated animals when compared to controls. Statistical analysis of the data did not reveal any significant intergroup differences. No obvious clinical signs of toxicity were detected for offspring from treated females when compared to controls.  The incidental clinical signs detected throughout the control and treated groups, consisting of small size, no milk in stomach, a physical injury, found dead or missing, were considered to be low incidence findings observed in offspring in studies of this type and were considered unrelated to test item toxicity.

 

In view of these results, the NOAEL ‘for systemic toxicity was considered to be 1000 mg/Kg bw/day The ‘No Observed Effect Level’ (NOEL) for reproductive/developmental toxicity was considered to be 1000 mg/Kg bw/day.

C9 members of the category

 

In a supporting Guideline (OECD 414) pre-natal developmental toxicity study (Harlan Laboratories, 2015), the test material (Nonene, branched; CAS# 97280-95-0) 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 100, 300, and 1000 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 surviving 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 weight, 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.

 

One female treated with 1000 mg/Kg bw/day was found dead on Day 16. This female was found to have a red stained ano-genital region and the bedding in the cage was also stained red. This death was most likely due to the intra-uterine total litter loss that was evident and, in the absence of any other significant effects in the remaining females or on fetus survival, it was considered unrelated to treatment. One female from this treatment group was killed in extremis on Day 8 due to a physical injury to one hind limb. This was due to a physical injury and unrelated to treatment.

 

The oral administration of the test material to pregnant rats by oral gavage during organogenesis at dose levels of 100, 300 and 1000 mg/Kg/day was associated with lower initial body weight gain and food intake in females treated with 1000 mg/Kg bw/day between Days 3 and 8 of gestation. Subsequently cumulative body weight gain throughout gestation and overall body weight gain when adjusted for the contribution of the gravid uterus were lower. However, body weight gain for these females during each measured period from Day 11 onwards was comparable to controls, therefore suggesting that the effect on body weight was an initial response to treatment and did not represent an overall adverse effect of treatment. Clinical signs were restricted to incidences of increased post-dosing salivation during the final two weeks of treatment.

 

There was no obvious adverse effect of maternal treatment on litter data as assessed by mean number of implantations, early and late embryonic/fetal deaths and live fetuses or sex ratio, as assessed by percentage male fetuses.

 

No treatment-related effects were detected on fetal external findings, skeletal development or in the type and incidence of skeletal or visceral findings. There were no findings that were considered to represent any known malformations.

 

Based on the results observed, the ‘No Observed Adverse Effect Level' (NOAEL) for the pregnant female was considered to be 1000 mg/Kg bw/day. No treatment-related changes were detected in the offspring parameters measured. The ‘No Observed Effect Level’ (NOEL) for developmental toxicity was therefore considered to be 1000 mg/Kg bw/day. 

 

In a Guideline (OECD 422) combined repeated dose reproductive/developmental toxicity study (Harlan Laboratories, 2014), the test material (Nonene, branched; CAS# 97280-95-0) was administered by gavage to three groups of Wistar Han™:RccHan™:WIST strain rats (12/sex/dose) for up to eight weeks (including a two week pre-pairing phase, pairing, gestation and early lactation for females), at dose levels of 100, 300 and 1000 mg/Kg bw/day. A control group of twelve males and twelve females was dosed with vehicle alone (Arachis oil BP).

 

Clinical signs, behavioral assessments, body weight change and food and water consumption were monitored during the study. Pairing of animals within each dose group was undertaken on a one male: one female basis within each treatment group on Day 15 of the study, with females subsequently being allowed to litter and rear their offspring to Day 5 of lactation. During the lactation phase, daily clinical observations were performed on all surviving offspring, together with litter size and offspring weights and assessment of surface righting reflex. Extensive functional observations were performed on five selected males from each dose group after the completion of the pairing phase, and for five selected parental females from each dose group on Day 4 post-partum. Hematology and blood chemistry were evaluated prior to termination on five selected males and females from each dose group.

 

Adult males were terminated on Day 43, followed by the termination of all females and offspring on Day 5 post-partum. Any female which did not produce a pregnancy was terminated on or after Day 25 post coitum. One female which did not show positive evidence of mating and did not produce a pregnancy was terminated on Day 57. All animals were subjected to a gross necropsy examination and histopathological evaluation of selected tissues was performed.

 

No test material related mortality was observed through the study period. Clinical signs were detected in animals of either sex treated with 300 and 1000 mg/Kg bw/day during the study. Episodes of increased salivation were reported from Day 8 onwards. The physical condition of males treated with 1000 mg/Kg bw/day was also affected with reductions in body weight development during Weeks 1 and 3 of treatment. Subsequently, a reduction in overall body weight gain in these males and a slight reduction in food efficiency during week 1, was evident. A reduction in haemoglobin was also observed in these males. Water consumption was also significantly increased for animals of either sex from all treatment groups throughout the treatment period. Observations of this nature are often reported when a test material formulation is unpalatable or irritant and can be associated with gastric irritancy rather than be attributable to systemic toxicity. This was supported microscopically with stomach changes identified as epithelial hyperplasia in the forestomach in animals of either sex treated with 1000 mg/Kg bw/day and in males treated with 300 mg/Kg bw/day. This finding was considered to be the result of local irritancy of the test material and therefore cannot be considered indicative of true systemic toxicity.

 

Although there were some statistically significant differences in treated animals from controls for the blood chemical parameters measured, these differences were considered not to be of toxicological significance. Macroscopic findings detected at necropsy were confined to enlarged, pale and mottled kidneys and a dark liver in a number of males treated with 1000 mg/Kg bw/day. One male treated with 300 mg/Kg bw/day also had enlarged kidneys.

 

Histopathological examination of the liver revealed hepatocellular hypertrophy in animals of either sex treated with 1000 and 300 mg/Kg bw/day and in males treated with 100 mg/Kg bw/day. Organ weight data supported this finding with increased absolute and relative liver weights observed in these animals. In the absence of any degenerative or inflammatory changes, this condition is considered to be adaptive in nature.

 

Microscopic examination of the thyroid revealed increased incidence and severity of follicular cell hypertrophy/hyperplasia in animals of either sex treated with 1000 and 300 mg/Kg bw/day and in males treated with 100 mg/Kg bw/day. Males treated with 1000 and 300 mg/Kg bw/day also showed hypertrophic/vacuolated cells in the pituitary. The thyroid, liver and pituitary changes are characteristic of a consequence of hepatocellular induction as a result of enhanced hepatic metabolism. As a side effect of hepatic induction an increased liver metabolism of thyroid hormones T3 and T4 can occur. This subsequently leads to an enhanced thyroid gland production of these hormones as a consequence of a negative feedback stimulation of TSH production. The appearance of thyroid follicular cell hypertrophy and hypertrophic/vacuolated cells in the pituitary are themselves considered to be a result of this process. The thyroid and pituitary changes were considered to be adaptive in nature.

 

Microscopic examination also revealed effects in the kidneys of males from all treatment groups. Tubular basophilia and hyaline droplets was present in males from all treated groups. These tubular findings were also accompanied by tubular degeneration/debris in males treated with 1000 and 300 mg/Kg bw/day. The hyaline droplets can be directly linked to accumulation of alpha 2u-globulin, which is unique to the male rat. This finding is commonly observed in male rats following treatment with some hydrocarbons and is not predictive of any adverse effect in humans. The remaining kidney findings consisting of tubular degeneration/debris may be considered to represent an adverse effect of the test item, however immunohistochemical staining demonstrated that these findings were correlated to the same condition as hyaline droplets.

 

Mating performance and fertility was unaffected by treatment. Offspring viability was however reduced in litters from females treated with 1000 mg/Kg bw/day on Day 4 post-partum. Subsequently reduced litter size and litter weights were evident in these litters on Day 4 post-partum when compared to controls. One female treated with 1000 mg/Kg bw/day also had a total litter loss between Days 2 and 4 post-partum. The mean offspring body weight gains for litters which survived to Day 5 post-partum were also reduced between Days 1 and 4 post-partum at this treatment level.

 

Based on the results observed, the No Observed Adverse Effect Level (NOAEL) for systemic toxicity was determined to be 1000 mg/Kg bw/day. The ‘No Observed Effect Level’ (NOEL) for reproductive toxicity was considered to be 300 mg/Kg bw/day, based on reduced offspring viability, offspring body weight gain, litter size and litter weights on Day 4 post-partum at 1000 mg/Kg bw/day.

C10 members of the category

 

In a supporting Guideline (OECD 422) combined repeated dose, reproductive/developmental toxicity screening test (Harlan Laboratories, 2014), the test material (Decene; CAS# 25339-53-1) was administered by gavage to three groups, each of twelve male and twelve female Wistar Han™:RccHan™:WIST strain rats, for up to eight weeks (including a two week pre-pairing phase, pairing, gestation and early lactation for females), at dose levels of 100, 300 and 1000 mg/Kg bw/day. A control group of twelve males and twelve females was dosed with vehicle alone (Arachis oil BP).

 

Clinical signs, behavioural assessments, body weight change and food and water consumptionwere monitored during the study.

 

Pairing of animals within each dose group was undertaken on a one male: one female basiswithin each treatment group on Day 15 of the study, with females subsequently being allowed tolitter and rear their offspring to Day 5 of lactation.

 

During the lactation phase, daily clinical observations were performed on all surviving offspring,together with litter size and offspring weights and assessment of surface righting reflex. Extensive functional observations were performed on five selected males from each dose group after the completion of the pairing phase, and for five selected parental females from each dose group on Day 4 post-partum. Haematology and blood chemistry were evaluated prior to termination on five selected males and females from each dose group.

 

Surviving adult males were terminated on Day 43, followed by the termination of all females andsurviving offspring on Day 5 post-partum. Any female which did not produce a pregnancy was terminated on or after Day 25 post coitum. Two females showing no visible vaginal opening were terminated on Day 42. All animals were subjected to a gross necropsy examination and histopathological evaluation of selected tissues was performed.

 

No test material related mortality was observed through the study period. Oral gavage administration of the test material at dosages of 100, 300 and 1000 mg/Kg bw/day for approximately six weeks resulted in minimal to moderate microscopic changes in the fore stomach of one male at 300 mg/Kg bw/day and both sexes at 1000 mg/Kg bw/day. These minimal to moderate histopathological changes in the fore stomach were considered to have occurred due to a local irritant effect, rather than true systemic toxicity. As a consequence, the NOAEL for adult systemic toxicity was considered to be 1000 mg/Kg bw/day.

 

There were no treatment-related effects on mating performance; fertility; or gestation length apparent at 100, 300 or 1000 mg/Kg bw/day. There was no effect of treatment on corpora lutea count, pre-implantation loss, numbers of implantations, post-implantation loss, litter size, sex ratio and subsequent offspring survival to Day 4 of age at 100, 300 or 1000 mg/Kg bw/day. Offspring body weight or body weight gain and litter weights, surface righting ability on Day 1 or clinical signs to Day 5 of age remained unaffected post exposure at 100, 300 or 1000 mg/Kg bw/day. Additionally, gross necropsy findings for offspring did not indicate any adverse effect of maternal treatment on offspring development at 100, 300 or 1000 mg/Kg bw/day.

 

Based on the results observed, the No Observed Effect Level (NOEL) for reproduction, including the survival, growth and development of the offspring, is considered to be 1000 mg/Kg bw/day.

C14 members of the category

 

In a supporting Guideline (OECD 422) combined repeated dose, reproductive/developmental toxicity study (Daniel, 1995), the test material (1-Tetradecene; CAS# 1120-36-1) was administered via oral gavage to Sprague-Dawley Crl:CD® BR VAF/Plus® rats (12 males and 20 females/dose) at doses of 0, 100, 500, or 1000 mg/Kg/day in corn oil. The F0 females were treated for 28 days prior to mating until the day prior to euthanasia (43-47 days). A satellite group of F0 females were added for neurotoxicity, clinical pathology and histopathological evaluations. Twelve F0 females assigned to a breeding phase were treated for 14 days prior to mating, during mating, gestation, and lactation until the day prior to euthanasia (42-51 days).

 

Animals were observed daily for overt signs of toxicity, and body weights and food consumption were measured at specific intervals. Breeding females were allowed to deliver and rear their offspring until lactation Day 4. Viability and development of the F1 generation were evaluated and surviving F1 pups were euthanized and necropsied on lactation Day 4. Functional observation battery, motor activity and clinical pathology evaluations were performed on selected F0 males and all satellite females. All F0 males and females were subjected to gross necropsy at the time of euthanasia. Specified tissues were retained at necropsy for selected F0 males and all (satellite and breeding) females and preserved in Bouin’s solution or 10% neutral buffered formalin. All tissues and organs from five randomly selected males and females from the control and high dose groups; lungs, liver, kidneys, and reproductive tract from all females; and gross lesions from all animals in each group were processed for microscopic examination.

 

Oral administration of the test material to virgin females and F0 parental animals produced minor clinical signs (salivation and urine staining) and dose-related hydrocarbon nephropathy in male rats in the 100, 500, and 1000 mg/Kg/day groups. This finding in male rats is commonly associated with hydrocarbon nephropathy and is not believed to pose any risk to humans.

 

In the satellite females, minor decreases were observed in the mean red cell count and hematocrit in 100, 500 and 1000 mg/Kg/day groups, as well as in haemoglobin and MCV in the 1000 mg/Kg/day group. Similar changes, though no statistically significant were observed in males. Therefore, the biological significance of these changes in the females is not clear.

 

In males, statistically significant minor increases in ALT were observed in 500 and 1000 mg/Kg/day groups. In satellite females, there were statistically significant decreases in sodium in 100, 500 and 1000 mg/Kg/day groups and increases in cholesterol in the 500 and 1000 mg/Kg/day groups. The changes of ALT sodium and cholesterol were observed in one sex and therefore were considered of no biological significance.

 

Minimal to moderate hepatocyte cytoplasmic vacuolation was observed in the livers of male and female rats at dosages of 500 and 1000 mg/Kg/day and correlated with increased liver weights at same tested doses. However, only the statistically significant increase of liver weight to brain weight ratio was seen in 1000 mg/Kg/day group females. The hepatocellular vacuolation was clearly related to the test item administration. However, in the absence of any other histopathological changes in liver as well as in the absence of significant effects on liver function (as indicated by the measured clinical chemistry parameters) the hepatocellular vacuolation might be considered to represent an adaptive response.

 

There were no other apparent test-article related histopathologic findings seen in any other tissues or organs examined. A statistically significant decrease in spleen weight relative to brain weight was noted in the 1000 mg/Kg/day group females. In addition, a statistically significant increase in absolute kidney weight was observed in 500 mg/Kg/day group females. These changes in female organs were not believed to be biologically meaningful because no test article-related lesions were seen histologically in these organs.

 

There was no evidence of impaired reproductive capabilities in the F0 generation or developmental toxicity in the F1 generation through lactation Day 4. The pup survival, litter sizes, sex ratios, external observations, body weights and necropsy of the F1 pups were comparable between all the groups.

 

Based on the results observed, the NOAEL for systemic toxicity was determined to be 100 mg/Kg/day in satellite females. Since hydrocarbon nephropathy was observed at all dose levels in male rats, a NOAEL for systemic toxicity could not be determined for males. The NOAEL for reproductive, developmental, or neurotoxicity was determined to be 1000 mg/Kg/day in male and female rats.

C16 members of the category

 

In a supporting Guideline (OECD 422) combined repeated dose, reproductive/developmental toxicity study (Harlan Laboratories, 2014), the test material (Hexadecene; CAS# 26952-14-7) was administered by gavage to three groups, each of twelve male and twelve female Wistar Han™:RccHan™:WIST strain rats, for approximately six weeks (including a two week pre-pairing phase, pairing, gestation and early lactation for females), at dose levels of 100, 300 and 1000 mg/Kg bw/day. A control group of twelve males and twelve females was dosed with vehicle alone (Arachis oil BP).

 

Clinical signs, behavioral assessments, body weight change and food and water consumption were monitored during the study.

 

Pairing of animals within each dose group was undertaken on a one male: one female basis within each treatment group on Day 15 of the study, with females subsequently being allowed to litter and rear their offspring to Day 5 of lactation.

 

During the lactation phase, daily clinical observations were performed on all surviving offspring, together with litter size and offspring weights and assessment of surface righting reflex. Extensive functional observations were performed on five selected males from each dose group after the completion of the pairing phase, and for five selected parental females from each dose group on Day 4 post-partum. Hematology and blood chemistry were evaluated prior to termination on five selected males and females from each dose group.

 

Adult males were terminated on Day 43, followed by the termination of all females and offspring on Day 5 post-partum. Any female which did not produce a pregnancy was terminated on or after Day 25 post coitum. One female which did not show positive evidence of mating and did not produce a pregnancy was terminated on Day 57. All animals were subjected to a gross necropsy examination and histopathological evaluation of selected tissues was performed.

 

No mortality was observed through the study period. Oral gavage administration of the test material at dosages up to 1000 mg/Kg bw/day for approximately six weeks (including two weeks pre-pairing, gestation and early lactation for females) was well tolerated, with no effect of treatment for body weight performance or food intake being apparent in either sex.

 

Treatment related clinical signs were restricted to increased post dosing salivation for both sexes at 1000 mg/Kg bw/day, and a few females at 300 mg/Kg bw/day but this was considered to reflect palatability of the test item rather than any systemic effect of treatment. Increased water consumption, compared to control, was apparent for males at 1000 mg/Kg bw/day, which may also reflect palatability of the test item.

 

Assessment of hematology and blood chemistry parameters did not reveal any differences considered to indicate an effect of treatment at dosages up to 1000 mg/Kg bw/day. Macroscopic necropsy findings and organ weights also did not indicate any treatment related effects.

 

Histopathological examination of tissues on the study did not reveal any findings that were considered to indicate systemic toxicity. At 1000 mg/Kg bw/day, minimal or moderate peribronchiolar inflammation was observed in the lungs for both sexes, but this was considered to be a consequence of accidental aspiration of the test item during the gavage procedure and to be of no toxicological significance.

 

There were no effects of treatment on the reproductive performance, including offspring survival, growth and development, at 100, 300 or 1000 mg/Kg bw/day.

 

Based on the results observed, 1000 mg/Kg bw/day was considered to represent the No Observed Effect Level (NOEL) for adult toxicity as well as a clear No Observed Adverse Effect Level (NOAEL).The No Observed Effect Level (NOEL) for reproduction, including the survival, growth and development of the offspring, is also considered to be 1000 mg/Kg bw/day.

C18 members of the category

 

In a key Guideline (OECD 414) pre-natal developmental toxicity study (Harlan Laboratories, 2015), the test material (Octadecene; CAS# 27070-58-2) was administered by gavage to three groups each of twenty-four time mated Sprague-Dawley Crl:CD® (SD) IGS BR strain rats, between Days 5 and 19 of gestation inclusive at dose levels 100, 300, and 1000 mg/Kg bw/day. A further group of twenty-four time mated females was exposed to the vehicle only (Arachis Oil BP) to serve as a control over the same treatment period.

 

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.

 

No mortality was observed through the study period and oral administration of the test material to pregnant rats by oral gavage during gestation at dose levels of 100, 300 and 1000 mg/Kg bw/day did not result in any treatment related effects. The No

Observed Effect Level' (NOEL) for the pregnant female was considered to be 1000 mg/Kg bw/day.

 

No treatment-related changes were detected in the offspring parameters measured. The ‘No Observed Effect Level’ (NOEL) for developmental toxicity was therefore considered to be 1000 mg/Kg bw/day. 

 

In a supporting Guideline (OECD 414) pre-natal developmental toxicity study (Harlan Laboratories, 2015), the test material (Octadec-1-ene; CAS# 112-88-9), 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 100, 300, and 1000 mg/Kg bw/day. A further group of twenty-four time mated females was exposed to the vehicle only (Arachis oil BP) 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 weight, 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.

 

No mortality was observed through the study period and the oral administration of the test material to pregnant rats by oral gavage during gestation days 3 to 19, at dose levels of 100, 300 and 1000 mg/Kg bw/day did not result in any treatment-related effects. The ‘No Observed Effect Level’ (NOEL) for the pregnant female was considered to be 1000 mg/Kg bw/day.

 

The number of implantations, subsequent embryofetal survival and litter size, sex ratio and mean fetal, litter and placental weights on Day 20 of gestation were unaffected by maternal treatment at 100, 300 and 1000 mg/Kg bw/day.

 

Neither the type, incidence or distribution of findings observed during external examination of the fetuses at necropsy on Day 20 of gestation and subsequent detailed visceral and skeletal examination indicated any adverse effect of maternal treatment on fetal development. Females treated with 1000 and 300 mg/Kg bw/day showed a statistically significant reduction in the percent of fetuses showing incomplete ossification of the frontal bone. Although this variant was statistically significant, it does not reflect a delay in development but rather a reduction in the number of fetuses and litters showing incomplete ossification of the frontal bone. There was no dose related response and the observation of one variant at a lower incidence compared with controls is not significant when evaluated in isolation. A true developmental effect is only seen when a number of variants or a syndrome of variants is observed, therefore the intergroup difference is considered of no biological significance.

 

No treatment-related changes were detected in the offspring parameters measured. The ‘No Observed Effect Level’ (NOEL) for developmental toxicity was therefore considered to be 1000 mg/Kg bw/day. 

 

In a supporting Guideline (OECD 422) repeated dose, reproductive/developmental toxicity screening study (Harlan Laboratories, 2014), the test material (Octadec-1-ene UVCB; CAS# 112-88-9) was administered by gavage to WistarHan™: RccHan™: WISTstrain rats (12/sex/dose), for up to eight weeks (including a two-week prepairing phase, pairing, gestation and early lactation for females), at dose levels of 100, 300 and 1000 mg/Kg bw/day. A control group of twelve males and twelve females was dosed with vehicle alone (Arachis oil BP).

 

Clinical signs, behavioral assessments, body weight change and food and water consumption were monitored during the study. Pairing of animals within each dose group was undertaken on a one male: one female basis within each treatment group on Day 15 of the study, with females subsequently being allowed to litter and rear their offspring to Day 5 of lactation. During the lactation phase, daily clinical observations were performed on all surviving offspring, together with litter size and offspring weights and assessment of surface righting reflex. Extensive functional observations were performed on five selected males from each dose group after the completion of the pairing phase, and for five selected parental females from each dose group on Day 4 post-partum. Hematology and blood chemistry were evaluated prior to termination on five selected males and females from each dose group. Adult males were terminated on Day 43, followed by the termination of all females and offspring on Day 5 postpartum. Any female which did not produce a pregnancy was terminated on or after Day 25 post coitum. All animals were subjected to a gross necropsy examination and histopathological evaluation of selected tissues was performed.

 

No mortality was observed through the study period. Clinical signs were confined to increased salivation detected in animals of either sex treated with 1000 and 300 mg/Kg bw/day. Observations of this nature are commonly observed following the oral administration of an unpalatable or slightly irritant test item formulation and do not necessarily represent an adverse effect of treatment.

 

No adverse effect was apparent for body weight or food consumption in treated animals throughout the treatment period. Although there were some statistically significant differences in treated animals from controls for the hematological and blood chemical parameters measured, these differences were considered not to be of toxicological significance. No macroscopic abnormalities were evident in treated animals; however microscopic examination revealed changes in the mesenteric lymph nodes and spleen. A minimal or mild inflammatory cell infiltrate was present in the periglandular fat surrounding the mesenteric lymph nodes in animals of either sex treated with 1000 and 300 mg/Kg bw/day. The infiltrate was mixed in character, principally composed of lymphocytes and neutrophils and tended to have a perivascular or perilymphatic orientation. Although the origin of the finding is uncertain it might have resulted from local irritation of material from the gut reaching the drainage nodes however there was no evidence of local irritation to the gut mucosa. However, in the absence of any other associated pathological changes at the low severity levels noted the change is considered likely to be non-adverse. The statement of laboratory is provided as an attachment to this IUCLID record.

The incidence and severity of extramedullary hematopoiesis was increased in the spleen in animals of either sex treated with 1000 mg/Kg bw/day and in females treated with 300 mg/Kg bw/day. The inter-group differences in females were not clearly dosage-related and taking into account the low number of animals examined, the absence of supportive intergroup differences in hematological values or bone marrow findings, these differences were considered more likely to have arisen as a result of individual variation, or possibly as a response to blood loss during parturition, than to be test item related. There were no statistically significant differences in adult organ weights that, in the absence of any evidence of histopathological change, were considered to be of any toxicological significance.

 

Mating performance and fertility was unaffected by treatment and there were no treatment-related effects detected in the offspring parameters observed.

 

Based on the results observed, the ‘No Observed Effect Level’ (NOEL) for systemic toxicity was considered to be 100 mg/Kg bw/day and the ‘No NOEL for reproductive/developmental toxicity was considered to be 1000 mg/Kg bw/day.

 

In an additional guideline (OECD 421) reproduction/developmental screening study (Thorsud, 2003), octadecenes, dissolved in corn oil, was administered to Sprague-Dawley rats (12 sex/dose) by gavage at dose levels of 0, 100, 500, or 1000 mg/Kg/day for 42 days.

 

Parents and offspring were observed for clinical signs of toxicity, mortality, mating, parturition, lactation, off-spring growth and maturity. Body weight and food consumption determinations were also conducted periodically through the study period. All parents and offspring were subjected to necroscopy following termination or death. 

 

There was no mortality observed in animals in the control, 100, 500, or 1000 mg/Kg/day dose group. No treatment-related or dose-dependant signs of clinical toxicity were noted in rats at any dose level. Mean body weight, body weight change and food consumption was observed to be normal in all treatment animals when compared with the controls. Parent female mating, fertility, and mean gestation lengths were observed to be comparable with controls as were the mean number of pups delivered and live birth and viability indices. Mean live pups/litter and sex ratio/litter were also found to be normal and comparable to those observed in control animals.

 

Gross necroscopy revealed no remarkable differences between octadecenes-treated and control animals. There were no microscopic lesions observed in male or female rats treated with octadecenes and no statistically significant differences in absolute or relative epididymides weight were noted in treated males when compared with control males. There were no statistically significant differences observed in pup weights on lactation days 1 and 4 and gross necroscopy on lactation day 4 revealed no treatment-related effects.

 

Based on the lack of adverse systemic effects observed in the study, the developmental/reproductive toxicity NOAEL for octadecene was reported as 1000 mg/Kg/day.

C12-30 members of the category

 

In a supporting Guideline (OECD 414) pre-natal developmental toxicity study (Envigo Research Ltd., 2016), the test material (Hydrocarbons, C12-30, olefin-rich, ethylene polymn by product CAS# 68911-05-7) was administered by gavage to three groups each of twenty-four time mated Sprague-Dawley Crl:CD® (SD) IGS BR strain rats, between Days 5 and 19 of gestation inclusive at dose levels 100, 300, and 1000 mg/Kg bw/day. A further group of twenty-four time mated females was exposed to the vehicle only (Arachis oil BP) 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.

 

No mortality was observed through the study period and oral administration of the test material to pregnant rats by oral gavage during gestation at dose levels of 100, 300 and 1000 mg/Kg bw/day did not result in any treatment related effects.

 

The ‘No Observed Effect Level’ (NOEL) for the pregnant female was considered to be 1000 mg/Kg bw/day. No treatment-related changes were detected in the offspring parameters measured. The NOEL for developmental toxicity was therefore considered to be 1000 mg/Kg bw/day.

No guideline developmental toxicity studies are available however in a 2nd species, which is a standard information requirement (REACH, Annex IX/X, section 8.7.2). Therefore, OECD 414 (2ndspecies – Rabbit) test proposals for the substances hex-1-ene, octadecene, octadec-1-ene, alkenes C8-10, C9-rich, and nonene, branched have been proposed. It is proposed that the studies will be carried out in rabbits using oral (gavage) administration and will follow the basic study design outlined in OECD Guideline 414.


Justification for selection of Effect on developmental toxicity: via oral route:
Oral screening level information is available on the potential of 5 members of this category covering C6 to C18 to cause developmental toxicity. No developmental effects were observed in female rats treated for the duration of pregnancy at dose levels up to 1000 mg/kg bw/day.

Justification for classification or non-classification

Guideline screening reproductive toxicity (OECD 421 and 422) and developmental toxicity (OECD 414) studies have been conducted for members of the Higher Olefins category covering C6 to C18.

 

The weight of evidence from oral reproductive and developmental toxicity studies, accompanied with data from oral and inhalation sub-chronic toxicity studies in rats indicate that category members have little or no potential to be considered reproductive/developmental toxicants.

 

Therefore, no classification is considered necessary under the current CLP regulation.