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Toxicological information

Toxicity to reproduction

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Endpoint:
screening for reproductive / developmental toxicity
Type of information:
read-across from supporting substance (structural analogue or surrogate)
Adequacy of study:
supporting study
Reliability:
2 (reliable with restrictions)
Justification for type of information:
1. HYPOTHESIS FOR THE ANALOGUE APPROACH
This read-across is based on the hypothesis that the target substance and 2 source substances have the same expected mode of action and similar physicochemical properties relevant for the read-across endpoints.
The justification of the proposed read-across to 2,6-dimethyl-5-heptenal and heptanoic acid is discussed in the following chapters.

2. SOURCE AND TARGET CHEMICAL(S) (INCLUDING INFORMATION ON PURITY AND IMPURITIES)
The target substance nonanal is a mono-constituent substance (EC 204-688-5, CAS 124-19-6). The typical concentration of the single constituent is 99.95%.
The source substance 2,6-dimethyl-5-heptenal (EC 203-427-2, CAS 106-72-9) is a mono-constituent substance. The typical concentration of the mono-constituents is 97.0%.
The source substance heptanoic acid (EC 203-838-7, CAS 111-14-8) is a mono-constituent substance. The typical concentration of the mono-constituents is 98.5%.
The chemical structure for the source substances is shown in Table 1 (RAAF Document) and the structural similarities for the source substances to undec-10-enal are given in Table 4 (RAAF Document).
The target substance and the source substances do not contain any impurities present at ≥ 1%. The purity of the test items within the respective REACH registration dossiers for 2-methylundecanal and heptanoic acid is > 99.00% and the data from the supplier Oxea Chemicals Ltd. for 2,6-dimethyl-5-heptenal indicates purity > 97.0% with no impurities > 1%.

3. ANALOGUE APPROACH JUSTIFICATION
The structures of the target and source substances are provided in Table 1. The target substance and the source substance have been characterised in this table using the categories and databases present in the OECD [Q]SAR Toolbox. From the profiling provided (Table 2), it can be seen that the 2 substances share structural similarities and also mechanistic actions which are both general and endpoint specific. This supports the hypothesis that the target and source substances have similar properties as a result of structural similarity and the same expected mode of action.
The OECD toolbox predicts all substances to be of low toxicity according to Cramer classes and both substances show no alerts according to DART Scheme v1.0.
Aldehyde C9 (nonanal) and 2,6-Dimethyl-5-heptenal are structurally similar aldehydes with a methyl substitution at the 2-position which is expected to be the primary site of metabolism. The primary route of metabolism for both substances is expected to be via rapid initial oxidation of the aldehyde function followed by glycine conjugation. This is supported by the most probable route of metabolism prediction of TIMES v.2.27.17 (rat in vivo model) as illustrated below.

4. DATA MATRIX
Please see the attached RAAF document.
Cross-referenceopen allclose all
Reason / purpose:
read-across source
Reference
Endpoint:
toxicity to reproduction
Remarks:
other: Screening test
Type of information:
experimental study
Adequacy of study:
weight of evidence
Study period:
No data
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: see 'Remark'
Remarks:
The study meets generally accepted scientific standards with acceptable restrictions for the standard test.
Justification for type of information:
1. HYPOTHESIS FOR THE ANALOGUE APPROACH
This read-across is based on the hypothesis that the target substance and 2 source substances have the same expected mode of action and similar physicochemical properties relevant for the read-across endpoints.
The justification of the proposed read-across to 2,6-dimethyl-5-heptenal and heptanoic acid is discussed in the following chapters.

2. SOURCE AND TARGET CHEMICAL(S) (INCLUDING INFORMATION ON PURITY AND IMPURITIES)
The target substance nonanal is a mono-constituent substance (EC 204-688-5, CAS 124-19-6). The typical concentration of the single constituent is 99.95%.
The source substance 2,6-dimethyl-5-heptenal (EC 203-427-2, CAS 106-72-9) is a mono-constituent substance. The typical concentration of the mono-constituents is 97.0%.
The source substance heptanoic acid (EC 203-838-7, CAS 111-14-8) is a mono-constituent substance. The typical concentration of the mono-constituents is 98.5%.
The target substance and the source substances do not contain any impurities present at ≥ 1%. The purity of the test items within the respective REACH registration dossiers for 2-methylundecanal and heptanoic acid is > 99.00% and the data from the supplier Oxea Chemicals Ltd. for 2,6-dimethyl-5-heptenal indicates purity > 97.0% with no impurities > 1%.

3. ANALOGUE APPROACH JUSTIFICATION
The structures of the target and source substances are provided in the RAAF document. The target substance and the source substance have been characterised in this table using the categories and databases present in the OECD [Q]SAR Toolbox. From the profiling provided (Table 2), it can be seen that the 2 substances share structural similarities and also mechanistic actions which are both general and endpoint specific. This supports the hypothesis that the target and source substances have similar properties as a result of structural similarity and the same expected mode of action.
The OECD toolbox predicts all substances to be of low toxicity according to Cramer classes and both substances show no alerts according to DART Scheme v1.0.
Aldehyde C9 (nonanal) and 2,6-Dimethyl-5-heptenal are structurally similar aldehydes with a methyl substitution at the 2-position which is expected to be the primary site of metabolism. The primary route of metabolism for both substances is expected to be via rapid initial oxidation of the aldehyde function followed by glycine conjugation. This is supported by the most probable route of metabolism prediction of TIMES v.2.27.17 (rat in vivo model) as illustrated below.
Qualifier:
according to
Guideline:
other: Unclear; makes reference to FDA (1987)
Deviations:
not specified
GLP compliance:
yes
Limit test:
yes
Species:
rat
Strain:
Sprague-Dawley
Sex:
female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: No data
- Age at study initiation: No data
- Weight at study initiation: No data
- Fasting period before study: No data
- Housing: No data
- Diet (e.g. ad libitum):No data
- Water (e.g. ad libitum):No data
- Acclimation period:No data
- Other: Nulliparous

ENVIRONMENTAL CONDITIONS
- Temperature (°C): No data
- Humidity (%): No data
- Air changes (per hr): No data
- Photoperiod (hrs dark / hrs light): No data
Route of administration:
oral: gavage
Vehicle:
corn oil
Details on exposure:
DIET PREPARATION
- Rate of preparation of diet (frequency): No data
- Mixing appropriate amounts with (Type of food):No data
- Storage temperature of food: No data

VEHICLE
- Justification for use and choice of vehicle: No data
- Concentration in vehicle: No data
- Amount of vehicle: 5 mL/kg bw/d
Analytical verification of doses or concentrations:
not specified
Duration of treatment / exposure:
1 week treatment followed by 7 d cohabitation period through gestation, parturition and 4-d postpartum period.
Frequency of treatment:
Daily
Remarks:
Doses / Concentrations:
300 mg/kg bw/d
Basis:
no data
Remarks:
Doses / Concentrations:
1500 mg/kg bw/d
Basis:
no data
Remarks:
Doses / Concentrations:
3000 mg/kg bw/d
Basis:
no data
No. of animals per sex per dose:
10 females per dose
Control animals:
yes, concurrent vehicle
Parental animals: Observations and examinations:
CAGE-SIDE OBSERVATIONS
- Viability was monitored twice daily during the study.
- Rats were observed daily for clinical signs approx. 30 mins after gavage administration.

BODY WEIGHT
- Measurement of body weight was performed weekly.

FOOD CONSUMPTION
- Food consumption measurement was also conducted weekly during the premating/premating period and then on days 0, 6, 14, 16, 21, and 25 of gestation and on days 1 and 4 of lactation/postparturition.

MATING PERFORMANCE
- Mating performance was evaluated daily during the cohabitation period.
- Dams were evaluated daily during gestation for duration of gestation, maternal behaviour, litter size and pup viability.

Oestrous cyclicity (parental animals):
Mating, day 0 of gestation identified on basis of spermatozoa in vaginal smear.
Postmortem examinations (parental animals):
SACRIFICE
- Dams that did not deliver litters were sacrificed on day 25 of presumed gestation and dams that did deliver litters were sacrificed on days 4 or 5 of lactation. All dams were examined for gross lesions and implantation sites.

GROSS NECROPSY
- Ovaries from all dams and any observed gross lesions were preserved in neutral 10% formalin for possible evaluation.
Postmortem examinations (offspring):
GROSS NECROPSY
- Vital signs at birth were determined for pups that were stillborn or died before the initial examination of the litter.
- Each litter was evaluated for viability a minimum of twice daily during the 4-day lactation period.
- Dead pups were removed and necropsied.
- Tissues with gross lesions were preserved for possible examination.
- Pups in each litter were counted and observed for nursing behaviour and physical abnormalities daily.
- Pup body weights were measured on days 1 and 4 of postpurition.
Statistics:
Fisher's ANOVA followed by Dunnett's test
Clinical signs:
effects observed, treatment-related
Dermal irritation (if dermal study):
not specified
Mortality:
mortality observed, treatment-related
Body weight and weight changes:
effects observed, treatment-related
Food consumption and compound intake (if feeding study):
effects observed, treatment-related
Food efficiency:
not specified
Water consumption and compound intake (if drinking water study):
not specified
Ophthalmological findings:
not examined
Haematological findings:
no effects observed
Clinical biochemistry findings:
not specified
Urinalysis findings:
not specified
Behaviour (functional findings):
not specified
Immunological findings:
not examined
Organ weight findings including organ / body weight ratios:
not examined
Gross pathological findings:
not examined
Neuropathological findings:
not examined
Histopathological findings: non-neoplastic:
not examined
Histopathological findings: neoplastic:
not specified
Other effects:
not specified
Reproductive function: oestrous cycle:
not examined
Reproductive function: sperm measures:
not examined
Reproductive performance:
not examined
CLINICAL SIGNS
- Clinical signs at 1500 and 3000 mg/kg in dams included decreased activity and excess salivation during the pregestation
period and increased (P<0.01) salivation in the high dose group during gestation.

BODY WEIGHT
- Significant (P<0.05 to <0.01) decreases in body weight and absolute and relative food consumption were measured during the premating period.
- Maternal body weights were decreased during gestation for the mid- and highdose groups of dams.
- Decreased body weights and absolute and relative food consumption in the 300 mg/kg bw/day group occurred only during premating and were not considered adverse effects.

MORTALITY
- 8/10 rats in the high dose group were moribund or found dead on days 2, 3,and 4 of the premating period.
- 1/2 surviving high-dose dams delivered a litter that died during the 4-day lactation period.

MATING AND FERTILITY
-Mating and fertility at the high dose were similar to controls.
-Measurements of mating success and fertility were similar for controls, low- and mid-dose groups.
Key result
Dose descriptor:
NOAEL
Effect level:
300 mg/kg bw/day
Based on:
test mat.
Sex:
female
Basis for effect level:
clinical signs
Key result
Dose descriptor:
LOAEL
Effect level:
1 500 mg/kg bw/day
Based on:
test mat.
Sex:
female
Basis for effect level:
clinical signs
Clinical signs:
effects observed, treatment-related
Dermal irritation (if dermal study):
no effects observed
Mortality / viability:
mortality observed, treatment-related
Body weight and weight changes:
effects observed, treatment-related
Food consumption and compound intake (if feeding study):
not specified
Food efficiency:
not specified
Water consumption and compound intake (if drinking water study):
not specified
Ophthalmological findings:
not specified
Haematological findings:
not specified
Clinical biochemistry findings:
not specified
Urinalysis findings:
not specified
Sexual maturation:
not examined
Organ weight findings including organ / body weight ratios:
not examined
Gross pathological findings:
no effects observed
Histopathological findings:
not examined
Other effects:
not specified
Behaviour (functional findings):
not specified
Developmental immunotoxicity:
not specified
VIABILITY
- Significant (P<0.05 to <0.01) decreases in pup viability occurred for middle and high dose groups as compared to controls.

NUMBER OF OFFSPRING
- The mid-dose litters were significantly less (P<0.05) than control group litters (sic).

BODY WEIGHT
- High-dose litters weighed remarkably less than controls.

OTHER
- No changes in averages for duration of cohabitation or gestation, implantation sites or pup sex ratios were seen at any dose levels.

GROSS PATHOLOGY
- No malformations or gross lesions in pups were attributable to the test material.
Key result
Dose descriptor:
LOAEL
Generation:
F1
Effect level:
1 500 mg/kg bw/day
Based on:
test mat.
Sex:
male/female
Basis for effect level:
clinical signs
Key result
Reproductive effects observed:
no
Lowest effective dose / conc.:
300 mg/kg bw/day
Treatment related:
no
Conclusions:
The test substance was assessed for toxicity to reproduction using a one-generation study in rats. The results showed that under the conditions of the test, dose levels of 300 mg/kg bw/day of the test material (5-heptenal, 2,6-dimethyl) had no adverse effects on the reproductive performance of female Sprague-Dawley rats or the growth or development of their offspring.
Reason / purpose:
read-across source
Reference
Endpoint:
one-generation reproductive toxicity
Type of information:
experimental study
Adequacy of study:
weight of evidence
Study period:
No data
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: see 'Remark'
Remarks:
It is a GLP compliant study conducted in compliance with agreed protocols, with no or minor deviations for the standard test.
Qualifier:
according to
Guideline:
other: Unclear; makes reference to FDA (1987)
Deviations:
not specified
GLP compliance:
yes
Limit test:
yes
Species:
rat
Strain:
Sprague-Dawley
Sex:
female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: No data
- Age at study initiation: No data
- Weight at study initiation: No data
- Fasting period before study: No data
- Housing: No data
- Diet (e.g. ad libitum):No data
- Water (e.g. ad libitum):No data
- Acclimation period:No data
- Other: Nulliparous

ENVIRONMENTAL CONDITIONS
- Temperature (°C): No data
- Humidity (%): No data
- Air changes (per hr): No data
- Photoperiod (hrs dark / hrs light): No data
Route of administration:
oral: gavage
Vehicle:
corn oil
Details on exposure:
DIET PREPARATION
- Rate of preparation of diet (frequency): No data
- Mixing appropriate amounts with (Type of food):No data
- Storage temperature of food: No data

VEHICLE
- Justification for use and choice of vehicle: No data
- Concentration in vehicle: No data
- Amount of vehicle: 5 mL/kg bw/d
Analytical verification of doses or concentrations:
not specified
Duration of treatment / exposure:
1 week treatment followed by 7 d cohabitation period through gestation, parturition and 4-d postpartum period.
Frequency of treatment:
Daily
Dose / conc.:
200 mg/kg bw/day
Dose / conc.:
1 000 mg/kg bw/day
Dose / conc.:
2 000 mg/kg bw/day
No. of animals per sex per dose:
10 females per dose
Control animals:
yes, concurrent vehicle
Parental animals: Observations and examinations:
CAGE-SIDE OBSERVATIONS
- Viability was monitored twice daily during the study.
- Rats were observed daily for clinical signs approx. 30 mins after gavage administration.

BODY WEIGHT
- Measurement of body weight was performed weekly.

FOOD CONSUMPTION
- Food consumption measurement was also conducted weekly during the premating/premating period and then on days 0, 6, 14, 16, 21, and 25 of gestation and on days 1 and 4 of lactation/postparturition.

MATING PERFORMANCE
- Mating performance was evaluated daily during the cohabitation period.
- Dams were evaluated daily during gestation for duration of gestation, maternal behaviour, litter size and pup viability.

Oestrous cyclicity (parental animals):
Mating, day 0 of gestation identified on basis of spermatozoa in vaginal smear.
Postmortem examinations (parental animals):
SACRIFICE
- Dams that did not deliver litters were sacrificed on day 25 of presumed gestation and dams that did deliver litters were sacrificed on days 4 or 5 of lactation. All dams were examined for gross lesions and implantation sites.

GROSS NECROPSY
- Ovaries from all dams and any observed gross lesions were preserved in neutral 10% formalin for possible evaluation.
Postmortem examinations (offspring):
GROSS NECROPSY
- Vital signs at birth were determined for pups that were stillborn or died before the initial examination of the litter.
- Each litter was evaluated for viability a minimum of twice daily during the 4-day lactation period.
- Dead pups were removed and necropsied.
- Tissues with gross lesions were preserved for possible examination.
- Pups in each litter were counted and observed for nursing behaviour and physical abnormalities daily.
- Pup body weights were measured on days 1 and 4 of postpurition.
Statistics:
Fisher's ANOVA followed by Dunnett's test
Clinical signs:
effects observed, treatment-related
Dermal irritation (if dermal study):
no effects observed
Mortality:
no mortality observed
Body weight and weight changes:
effects observed, treatment-related
Food consumption and compound intake (if feeding study):
effects observed, treatment-related
Food efficiency:
effects observed, treatment-related
Water consumption and compound intake (if drinking water study):
no effects observed
Ophthalmological findings:
not specified
Haematological findings:
not specified
Clinical biochemistry findings:
not specified
Urinalysis findings:
not specified
Behaviour (functional findings):
not specified
Immunological findings:
not specified
Organ weight findings including organ / body weight ratios:
not examined
Gross pathological findings:
not examined
Neuropathological findings:
not specified
Histopathological findings: non-neoplastic:
not examined
Histopathological findings: neoplastic:
not specified
Other effects:
not specified
Reproductive function: oestrous cycle:
not examined
Reproductive function: sperm measures:
not examined
Reproductive performance:
not examined
MORTALITY
- 1 and 3 deaths were reported in the 1000 and 2000 mg/kg bw/day dose groups, respectively.

CLINICAL SIGNS
- Clinical signs at 200 mg/kg bw/day in dams during premating and gestation included a significant increase in rales (P<0.01). This effect was not
reported during the lactation period. In the 1000 and 2000 mg/kg bw/day dose group, significant increases in the incidence of rales (P<0.01), excess salivation (P<0.01) was reported during premating and gestation. Excess salivation continued during lactation in the high-dose group.
- Other significant (P<0.01) effects during gestation in the high-dose group included decreased activity, ungroomed coat and labored breathing.

BODY WEIGHTS
- The 2000 mg/kg bw/day group showed reduced body weight gains during premating, and significantly (P<0.05 to <0.01) decreased average maternal body weights on days 10 and 16 of gestation.

FOOD CONSUMPTION
- Average and relative food consumption was reduced in the high-dose group of dams throughout the study. The high dose also was associated with reduced mating and fertility that were related to mortality.

COHABITATION INDICES
-The duration of cohabitation and fertility and gestation indices 200, 1000, or 2000 mg/kg bw/day were not different from comparable indices in thee control group.

Key result
Dose descriptor:
NOAEL
Effect level:
200 mg/kg bw/day
Based on:
test mat.
Sex:
female
Basis for effect level:
clinical signs
Key result
Dose descriptor:
LOAEL
Effect level:
1 500 mg/kg bw/day
Based on:
test mat.
Sex:
female
Basis for effect level:
clinical signs
Clinical signs:
no effects observed
Dermal irritation (if dermal study):
no effects observed
Mortality / viability:
no mortality observed
Body weight and weight changes:
effects observed, treatment-related
Food consumption and compound intake (if feeding study):
effects observed, non-treatment-related
Food efficiency:
not specified
Water consumption and compound intake (if drinking water study):
not specified
Ophthalmological findings:
not specified
Haematological findings:
not specified
Clinical biochemistry findings:
not specified
Urinalysis findings:
not specified
Sexual maturation:
not examined
Organ weight findings including organ / body weight ratios:
not examined
Gross pathological findings:
no effects observed
Histopathological findings:
not examined
Other effects:
not specified
Behaviour (functional findings):
not specified
Developmental immunotoxicity:
not specified
BODY WEIGHT
- The high-dose group exhibited reduced pup weights on day 4 postparturition.

OTHER
No biologically relevant or statistically significant differences in the number of implantations, duration of gestation, the percentage of dams delivering one or more live pups, and the pup viability index were observed.

GROSS PATHOLOGY
- No malformations or gross lesions were observed in pups at any dose levels.
Key result
Dose descriptor:
NOEL
Generation:
F1
Effect level:
200 mg/kg bw/day
Based on:
test mat.
Sex:
male/female
Basis for effect level:
clinical signs
Reproductive effects observed:
not specified
Conclusions:
The test substance was assessed for toxicity to reproduction using a one-generation study in rats. The results showed that under the conditions of the test, dose levels of 200 mg/kg bw/day of the test substance had no significant adverse effects on the reproductive performance of female Sprague-Dawley rats or the growth or development of their offspring.

Data source

Materials and methods

Results and discussion

Applicant's summary and conclusion

Conclusions:
The test substance was assessed for toxicity to reproduction using a one-generation study in rats using the analogue substances 2,6-dimethylhept-5-enal and Heptanoic acid. The results showed that under the conditions of the test, dose levels of 300 mg/kg bw/day of 2,6-dimethylhept-5 -enal and dose levels of 200 mg/kg bw/day of Heptanoic acid had no adverse effects on the reproductive performance of female Sprague-Dawley rats or the growth or development of their offspring.

The read-across are considered to be suitable based on the structural and “mechanistic action” similarities between the target substance (Nonanal) and source substances (2,6-dimethylhept-5-enal and Heptanoic acid) and their similar physico-chemical properties.