2,6-dimethyloctan-2-ol

Endpoint summary

Key value for chemical safety assessment

Effects on fertility

Description of key information

Toxicity to reproduction (OECDTG422): NOAEL >= 868.7 mg/kg bw/day for males, >=954.5 mg/kg bw/day for females (read-across)

Link to relevant study records

Reference

Reference 1

Endpoint:

screening for reproductive / developmental toxicity

Type of information:

experimental study

Adequacy of study:

key study

Study period:

25 March 2015 - 27 November 2015

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Justification for type of information:

This information is used for read across to Tetrahydromyrcenol.

Reason / purpose for cross-reference:

read-across: supporting information

Qualifier:

according to guideline

Guideline:

OECD Guideline 422 (Combined Repeated Dose Toxicity Study with the Reproduction / Developmental Toxicity Screening Test)

Version / remarks:

22 March 1996

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Limit test:

no

Species:

rat

Strain:

other: Crl:CD(SD)

Details on species / strain selection:

The rat was chosen as the test species because of the requirement for a rodent species by regulatory agencies.
The Crl:CD(SD) strain was used because of the historical control data available at this laboratory.

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Charles River (UK) Ltd.
- Females nulliparous and non-pregnant: Yes
- Age at study initiation: Approx. 10 weeks
- Weight at study initiation: Males: 335-402 g / Females: 230-291 g
- Fasting period before study: Only fasted overnight before blood sampling for haematology and blood chemistry investigations
- Housing: Polycarbonate body with a stainless steel mesh lid (changed at appropriate intervals) used during the acclimatisation, pre-pairing, gestation, littering and lactation periods. Polypropylene cages used during pairing.
- Diet: Ad libitum (but removed overnight before blood sampling for haematology and blood chemistry investigations)
- Water (e.g. ad libitum): Ad libitum (but removed overnight during urine collection)
- Acclimation period: Six days before start of treatment

DETAILS OF FOOD AND WATER QUALITY: SDS VRF1 Certified powdered diet; Potable water from the public supply via polycarbonate bottles with sipper tubes. Bottles were changed at appropriate intervals.

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 19-23
- Humidity (%): 40-70
- Air supply: Filtered fresh air which was passed to atmosphere and not recirculated
- Photoperiod (hrs dark / hrs light): 12/12

IN-LIFE DATES:
Animal arrival: 24 June 2015
F0 necropsy: Toxicity phase 13 August 2015, Recovery phase 28 August 2015, Reproductive phase
19 to 24 August 2015

Route of administration:

oral: feed

Vehicle:

corn oil

Remarks:

used as stabiliser

Details on exposure:

DIET PREPARATION
- Rate of preparation of diet (frequency): Weekly
- Mixing appropriate amounts with (Type of food): SDS VRF1 certified powdered diet
- Storage temperature of food: Ambient temperature (nominally 21°C) for eight days. Deep-frozen (nominally -20°C) for 22 days.

VEHICLE
- Justification for use and choice of vehicle (if other than water): The dietary of administration was chosen to simulate a condition of potential human exposure.
- Concentration in vehicle: test material to corn oil ratio 5:1

Details on mating procedure:

- M/F ratio per cage: 1:1 from within the same treatment groups.
- Length of cohabitation: Up to two weeks.
- Proof of pregnancy: Ejected copulation plugs in cage tray or sperm in the vaginal smear referred to as day 0 of gestation
- After successful mating each pregnant female was caged: individually

Analytical verification of doses or concentrations:

yes

Details on analytical verification of doses or concentrations:

For Week 1 and Final Week of treatment, representative samples of test diet were taken and submitted for analysis using chromatography. Each diet sample was sub-sampled (10 g ± 0.5 g) in duplicate and analysed in accordance with the analytical procedure. The analysed concentrations of 2,6-dimethyloct-7-en-2-ol in the test formulations analysed during the study, associated procedural recovery data and the deviation of mean results from nominal values were between -6% and +3.3% which is within applied limits of +10%/-15%. This confirms the accuracy of formulation. Procedural recovery values remained within 99.2% and 100.7%, confirming the continued accuracy of the method. The difference of individual values from the mean was <5% confirming the precision of analysis.

Duration of treatment / exposure:

Reproductive phase females: Three weeks before pairing, then throughout pairing and gestation until Day 6 of lactation.
Toxicity phase males: Three weeks pre-pairing up to necropsy after minimum of five weeks.

Frequency of treatment:

Continuous

Details on study schedule:

- Age at mating of the mated animals in the study: 13 weeks

Dose / conc.:

15 000 ppm

Remarks:

Corresponding actual intake:
Week 1 to 3 prior to pairing: 954.5 mg/kg bw/day
Days 0-19 of gestation: 975.8 mg/kg bw/day
Days 1-6 of lactation: 1449.4 mg/kg bw/day
Males: 868.7 mg/kg bw/day

Dose / conc.:

5 000 ppm

Remarks:

Corresponding actual intake:
Week 1 to 3 prior to pairing: 349.8 mg/kg bw/day
Days 0-19 of gestation: 342.7 mg/kg bw/day
Days 1-6 of lactation: 599.2 mg/kg bw/day
Males: 291.7 mg/kg bw/day

Dose / conc.:

1 500 ppm

Remarks:

Corresponding actual intake
Week 1 to 3 prior to pairing: 108 mg/kg/day
Days 0-19 of gestation: 102.3 mg/kg/day
Days 1-6 of lactation: 202.4 mg/kg/day
Males: 89 mg/kg bw/day

No. of animals per sex per dose:

10 females per dose group
5 males for control and high-dose group
10 males for low and mid-dose group

Control animals:

yes, concurrent vehicle

yes, historical

Details on study design:

- Dose selection rationale: The dietary levels of 1500, 5000 and 15000 ppm were selected based on the results of a 21-day preliminary study (Envigo Study Number: CIZ0001). In that study, administration of 2,6-dimethyloct-7-en-2-ol to CD rats by dietary administration for 21 days at dietary levels of 1500, 7500 or 15000 ppm was well tolerated. No animals died and the clinical condition of the animals was satisfactory.
- Rationale for animal assignment: random (Note: On Day 1 of study all animals were weighed and body weights were reviewed before dosing commenced by Study Management. The groups were adjusted to reduce inter-/intra-group variation.)

Positive control:

Not included

Parental animals: Observations and examinations:

CAGE SIDE OBSERVATIONS: Yes
- Time schedule: Twice daily

DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: Daily

BODY WEIGHT: Yes
Reproductive phase females: Before feeding on the day that treatment commenced (Week 0) and weekly before pairing, Days 0, 6, 13 and 20 after mating, Day 1, 4, and 7 of lactation and on the day of necropsy.

OTHER EXAMINATIONS: During the study, clinical condition, detailed physical examination and arena observations, sensory reactivity, grip strength, motor activity, body weight, food consumption, ophthalmic examination, haematology (peripheral blood), blood chemistry, urinalysis, oestrous cycles, pre-coital interval, mating performance, fertility, gestation length, organ weight and macroscopic pathology and histopathology investigations were undertaken.

Oestrous cyclicity (parental animals):

Dry smears: for 15 days before pairing using cotton swabs.
Wet smears: after pairing until mating, using pipette lavage.
Group 2 female 70: for five days after suspected mating using pipette lavage.

The following classifications of oestrous cycles is used:
Regular: All observed cycles of 4 or 5 days
Irregular: At least one cycle of 2, 3 or 6 to 10 days
Acyclic: At least 10 days without oestrus

Sperm parameters (parental animals):

Parameters examined in Parental male generation:
- testis weight
- tubular stages of the spermatogenic cycle
- missing germ cell layers or types
- retained spermatids
- multinucleate or apoptotic germ cells
- sloughing of spermatogenic cells in the lumen
Any cell- or stage-specificity of testicular findings was noted.

Litter observations:

PARAMETERS EXAMINED
The following parameters were examined in [F1] offspring:
- Clinical observations: were examined at approximately 24 hours after birth (Day 1 of age) and then daily thereafter for evidence of ill health or reaction to treatment; these were on an individual offspring basis or for the litter as a whole, as appropriate.
- Litter size: Daily records were maintained of mortality and consequent changes in litter size from Days 1-7 of age.
- Sex ratio of each litter: Recorded on Days 1, 4 and 7 of age.
- Individual offspring body weights: Days 1, 4 and 7 of age.
- Macropathology: for all offspring were also assessed.

GROSS EXAMINATION OF DEAD PUPS:
yes, where possible, a fresh macroscopic examination (external and internal) with an assessment of stomach for milk content was performed.

Postmortem examinations (parental animals):

SACRIFICE
- Male animals: All surviving animals after the Week 5 investigations completed.
- Maternal animals: All surviving animals at Day 7 of lactation.

GROSS NECROPSY
- Gross necropsy consisted of external and internal examinations

HISTOPATHOLOGY / ORGAN WEIGHTS
Pathology procedures (weiging, fixing histolgy and pathology) was performed for the five lowest numbered surviving toxicity phase males and all toxicity phase females and recovery phase animals at scheduled termination (described in the repeated dose toxicity endpoint summary).
For remaining toxicity males and reproductive phase females, the tissues indicated inbelow were prepared for microscopic examination and weighed, respectively:
- Epididymides
- Ovaries
- Prostate
- Seminal vesicles
- Testes
- Uterus
- Vagina

Postmortem examinations (offspring):

SACRIFICE
- The F1 offspring were sacrificed at day 7 of lactation.
- These animals were subjected to postmortem examinations

GROSS NECROPSY
- Where possible, a fresh macroscopic examination (external and internal) with an assessment of stomach for milk content was performed

Statistics:

All statistical analyses were carried out separately for males and females. For all other adult parameters, the analyses were carried out using the individual animal as the basic experimental unit. For litter/fetal findings the litter was taken as the treated unit and the basis for statistical analysis and biological significance was assessed with relevance to the severity of the anomaly and the incidence of the finding within the background control population.

The following sequence of statistical tests were used for grip strength, motor activity, body weight, food consumption, clinical pathology, litter size and survival indices, organ weight data:
- Bartlett's test for variance homogeneity (Bartlett 1937), followed by
Parametric analysis:
- Analysis of variance for any group differences
- Inter group comparisons using t-tests, error mean square from one-way analysis of variance
- F1 approximate test
- Williams' test for a monotonic trend
- Dunnett's test (Dunnett 1955, 1964) for non-monotonic trend
Non-parametric analysis
- Logarithmic and square-root transformations
- Kruskal-Wallis’ test (Kruskal and Wallis 1952, 1953) for group differences
- Inter group comparisons using Wilcoxon rank sum tests (Wilcoxon 1945)
- H1 approximate test for monotonicity of dose-response
- Shirley's test for a monotonic trend
- Steel's test (Steel 1959) for non-monotonic trend
For grip strength, motor activity, clinical pathology, litter size and survival indices:
- Fisher’s Exact tests (Fisher 1973)
- Pairwise comparisons for treatment groups against control group
- Generalised mixed linear model with binomial errors, a logit link function and litter as a random effect (Lipsitz 1991) for sex ratio
- Treatment group comparison to control using a Wald chi-square test
- Exact two-tailed Linear-by-linear test (Cytel 1995) for gestation length (asymptotic version used if needed)
For organ weight data:
- Analysis of covariance (Angervall and Carlstrom, 1963) for parametric methods

Reproductive indices:

Percentage mating (%) = Number animals mating / Animals paired x 100
Conception rate (%) = Number animals achieving pregnancy / Animals mated x 100
Fertility index (%) = Number animals achieving pregnancy / Animals pairing x 100
Gestation index (%) = Number of live litters born / number pregnant x 100

Offspring viability indices:

Post-implantation survival index (%) = Total number of offspring born / Total number of uterine implantation sites x 100
Live birth index (%) = Number of live offspring on Day 1 after littering / Total number of offspring born x 100
Viability index (%) = Number of live offspring on Day 7 / Number of liver offspring on Day 1 after littering x 100

Clinical signs:

no effects observed

Description (incidence and severity):

There were no signs observed, throughout the study, that were considered to be attributable to treatment with 2,6-dimethyloct-7-en-2-ol.

Mortality:

no mortality observed

Description (incidence):

No mortalities occured related to treatment with 2,6-dimethyloct-7-en-2-ol.

Body weight and weight changes:

effects observed, treatment-related

Description (incidence and severity):

Males: Unaffected by treatment
Reproductive phase Females: body weight gain was significantly reduced from Day 13-20 of gestation in females treated at 5000 or 15000 ppm.
Lactation phase Females: Day 1-7: body weight gain was significantly reduced in females treated at 15000 ppm.

Food consumption and compound intake (if feeding study):

effects observed, treatment-related

Description (incidence and severity):

During gestation, food intake in females treated at 15000 ppm food intake remained slightly low when compared with controls, from Day 0 to Day 3; thereafter food intake improved and was similar to that of controls.
During lactation, food intake was slightly low in females treated at 5000 ppm with a markedly low food intake observed in females treated at 15000 ppm. This effect persisted throughout lactation (until Day 7).

Food efficiency:

not specified

Water consumption and compound intake (if drinking water study):

effects observed, treatment-related

Description (incidence and severity):

On occasions throughout the treatment period, animals treated at 15000 ppm were observed to consume more water than the animals treated at 0, 1500 or 5000 ppm.
This effect is considered to be due to the palatability of the test material and not an adverse response to treatment.

Ophthalmological findings:

no effects observed

Description (incidence and severity):

Ophthalmic examination revealed no findings that could be attributed to treatment with 2,6- dimethyloct-7-en-2-ol.

Haematological findings:

effects observed, treatment-related

Description (incidence and severity):

None of the findings were conclusively linked to any of the treatment related microscopic pathology changes, and since all of the affected parameters evaluated during Week 2 of recovery showed recovery and, the survival, clinical condition and reproductive performance of the animals was unaffected by treatment, they are considered not to represent an adverse effect of treatment.

Males 15000 ppm:
- Significant increases in mean cell haemoglobin (p<0.05) and consequently mean cell haemoglobin concentration (p<0.01);
- Significant reduction in platelet counts (p<0.05)

Females 5000 or 15000 ppm:
- Mean cell haemoglobin concentration was significantly increased (p<0.05 in both groups)
- Mean cell volume was significantly reduced in females treated at 15000 ppm.

Clinical biochemistry findings:

effects observed, treatment-related

Description (incidence and severity):

None of the findings were conclusively linked to any of the treatment related microscopic pathology changes, and since all of the affected parameters evaluated during Week 2 of recovery showed recovery and, the survival, clinical condition and reproductive performance of the animals was unaffected by treatment, they are considered not to represent an adverse effect of treatment.

Males 15000 ppm:
- glucose concentration significantly reduced (p<0.05)
- chloride concentration significantly reduced (p<0.05)
- albumin/globulin ratio significantly reduced (p<0.01)

Females 1500, 5000 or 15000 ppm:
- glucose concentration significantly reduced (p<0.05, p<0.01 and p<0.01 respectively)
- potassium concentration significantly reduced (5000 or 15000 ppm) (p<0.01 both)
- albumin/globulin ratio reduced (15000 ppm)
- cholesterol increased (15000 ppm)

Urinalysis findings:

effects observed, treatment-related

Description (incidence and severity):

None of the findings were conclusively linked to any of the treatment related microscopic pathology changes, and since all of the affected parameters evaluated during Week 2 of recovery showed recovery and, the survival, clinical condition and reproductive performance of the animals was unaffected by treatment, they are considered not to represent an adverse effect of treatment.

Males 15000 ppm:
- low pH (not significant)
- significantly increased specific gravity (p<0.05)

Females 15000 ppm:
- increase in urinary volume (not significant)
- significant increase in total potassium (p<0.05)
- significant increase in total chloride levels (p<0.01)

Behaviour (functional findings):

no effects observed

Immunological findings:

not specified

Organ weight findings including organ / body weight ratios:

effects observed, treatment-related

Histopathological findings: non-neoplastic:

effects observed, treatment-related

Description (incidence and severity):

Changes related to treatment with 2,6-dimethyloct-7-en-2-ol were seen in the kidneys, liver and female reproductive tract.

Males:
- Kidneys: Hyaline droplets in the cortical tubular epithelium were evident in the kidneys from all males treated at 15000 ppm examined histopathologically and one male treated at 5000 ppm. In the most severely affected animal this change was accompanied by slight multifocal cortical tubular basophilia and minimal granular casts. Note: The accumulation of hyaline droplets in the cortical tubular epithelium of the kidneys, as sociated with granular casts and tubular basophilia is consistent with the finding of alpha 2u-globulin nephropathy (Frazier et al., 2012). This is a male rat specific finding associated with the binding of xenobiotics to this protein and interference with the normal lysosomal uptake and degradation of the protein. There was evidence of recovery from this change following 2 weeks respite from treatment.
- Liver: Portal pigment was seen in the bile ducts of three males receiving 15000 ppm. These consisted of deposits of brown pigment, predominantly located in the lumen of smaller bile ductules. Application of special stains to additional sections of liver showed this pigment to be Schmorl’s, Perls’ and Fouchet’s negative. Note: In the absence of any effect on the survival or reproductive performance of the animals and, no indication of impaired liver function from the blood chemistry analyses, these findings were cons idered not to represent an adverse effect of treatment.

Histopathological findings: neoplastic:

no effects observed

Other effects:

no effects observed

Reproductive function: oestrous cycle:

effects observed, treatment-related

Description (incidence and severity):

Female reproductive tract:
- All females receiving 15000 ppm were in diestrus, whereas Control females and females receiving 1500 or 5000 ppm were more often in proestrus or metoestrus. Uterine luminal dilatation was not evident in any of the females receiving 15000 ppm. The apparent difference in stage of oestrus is of uncertain significance. Note: This finding may be related to the test article related bodyweight effects seen in females of this dose group during treatment, but was considered to be of uncertain relationship to treatment since the efect was reversible and the oestrus cycles prior to pairing, mating performance and fertility of these females had been unaffected by treatment.

Reproductive function: sperm measures:

no effects observed

Reproductive performance:

no effects observed

Key result

Dose descriptor:

NOAEL

Remarks:

reproduction

Effect level:

>= 868.7 mg/kg bw/day (actual dose received)

Based on:

test mat.

Sex:

male

Basis for effect level:

other: NOAEL = Highest dose tested

Key result

Dose descriptor:

NOAEL

Remarks:

pre-mating

Effect level:

>= 954.5 mg/kg bw/day (actual dose received)

Based on:

test mat.

Sex:

female

Basis for effect level:

other: NOAEL = highest dose tested

Dose descriptor:

NOAEL

Remarks:

gestation

Effect level:

>= 975.8 mg/kg bw/day (actual dose received)

Based on:

test mat.

Sex:

female

Basis for effect level:

other: NOAEL = Highest dose tested

Dose descriptor:

NOAEL

Remarks:

lactation

Effect level:

>= 1 449.4 mg/kg bw/day (actual dose received)

Based on:

test mat.

Sex:

female

Basis for effect level:

other: NOAEL = Highest dose tested

Key result

Critical effects observed:

no

Clinical signs:

no effects observed

Mortality / viability:

mortality observed, non-treatment-related

Description (incidence and severity):

Mortality was observed in litters of all test groups (including control), but this is not considered treatment related.

Body weight and weight changes:

effects observed, treatment-related

Description (incidence and severity):

There was no difference in offspring body weight gain from Day 1-4 of age when compared with Controls; growth thereafter (Day 4-7) was lower in male and female offspring from parents receiving 15000 ppm. There was no effect of treatment in offspring from parents receiving 5000 or 1500 ppm.

The lower growth in the offspring of females treated at 15000 ppm, (Day 1-7) is likeley cased by the low food intake of the P0 females during this period. This low growth of the offspring was considered not to represent an adverse effect of treatment since the survival, clinical condition and macropathology findings for the offspring were not affected by treatment.

Food consumption and compound intake (if feeding study):

no effects observed

Description (incidence and severity):

Animals of the F1 generation were not dosed, any exposure was in utero or via the milk.

Food efficiency:

not specified

Water consumption and compound intake (if drinking water study):

no effects observed

Ophthalmological findings:

not examined

Haematological findings:

not examined

Clinical biochemistry findings:

not examined

Urinalysis findings:

not examined

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:

no effects observed

Behaviour (functional findings):

not examined

Developmental immunotoxicity:

not examined

Key result

Dose descriptor:

NOAEL

Remarks:

developmental (gestation)

Generation:

F1

Effect level:

>= 975.8 mg/kg bw/day

Based on:

test mat.

Remarks:

received via dams

Sex:

male/female

Basis for effect level:

other: NOAEL = Maximum tested dose level

Dose descriptor:

NOAEL

Remarks:

developmental (lactation)

Generation:

F1

Effect level:

>= 1 449.4 mg/kg bw/day

Based on:

test mat.

Remarks:

received via dams

Sex:

male/female

Basis for effect level:

other: NOAEL = Maximum tested dose level

Key result

Critical effects observed:

no

Key result

Reproductive effects observed:

no

Minor treatment related effects were observed for body weight, histopathology, blood chemistry and urine analysis in the parental animals. However, as these showed recovery and neither the survival nor the clinical condition were affected by treatment,they are considered not to represent an adverse effect of the treatment. The alpha 2u-globulin nephropathy observed in the kidneys of the male rat is a male-rat-specific finding associated with the binding of xenobiotics to this protein and interference with the normal lysosomal uptake and degradation of the protein. The histological changes observed in the liver were also not considered adverse, as no effect on the survival or reproductive performance of the animals was observed and no indication of impaired liver function could be seen from the blood chemistry analyses.

Conclusions:

Under the conditions of this study, the NOAEL for reproductive toxicity was determined to be at >=15000 ppm, representing mean achieved doses of 868.7 mg/kg bw/day.

Executive summary:

Dihdyromyrcenol is tested in an OECD TG 422 test. The repeated dose toxicity is repeated here and includes the developmental effects to presente a complete picture. In the present executive summary the reproductive/developmental toxicity effects are reported following dietary administration. A similarly constituted control group was assigned, and received the vehicle, powdered SDS VRF1 certified diet with corn oil, throughout the same relative treatment period.

Reproductive females were treated daily for three weeks before pairing, throughout pairing, gestation and until Day 6 of lactation. Females were allowed to litter and rear their offspring to weaning and were killed on Day 7 of lactation. The F1 generation were killed on Day 7 of age, but did not directly receive the test substance; any exposure was in utero or via the milk. Toxicity phase males were treated daily for three weeks before pairing up to necropsy after a minimum of five consecutive weeks.

During the study, clinical condition, detailed physical examination and arena observations, sensory reactivity, grip strength, motor activity, body weight, food consumption, ophthalmic examination, haematology (peripheral blood), blood chemistry, urinalysis, oestrous cycles, pre-coital interval, mating performance, fertility, gestation length, organ weight and macroscopic pathology and histopathology investigations were undertaken. The clinical condition, litter size and survival, sex ratio,bodyweightand macropathology for all offspring were also assessed.

Results

Clinical signs: There were no adverse effects observed attributed to treatment on: clinical condition, sensory reaction, grip strength, motor activity, ophthalmic examination and macroscopic pathology.  

Other effects: Minor treatment related effects were observed for body weight, histopathology, blood chemistry and urine analysis. However, the animals showed recovery and, the survival, clinical condition and reproductive performance of the animals was unaffected by treatment, therefore they are considered not to represent an adverse effect of the treatment. In the kidney of male rats, alpha 2u-globulin nephropathy was observed. This is a male-rat-specific finding associated with the binding of xenobiotics to this protein and interference with the normal lysosomal uptake and degradation of the protein, and did not adversly affect the reproductive performance of the animals. Some histological changes were observed in the liver (portal pigment), also these effect were not considered adverse, as no effect on the survival or reproductive performance of the animals was observed and no indication of impaired liver function could be seen from the blood chemistry analyses.

Fertility effects: In the female reproductive tract, all females receiving 15000 ppm were observed to be in diestrus in comparison to the Controls and low and intermediate dosage groups. This was associated with a reduced incidence of luminal dilatation of the uterus, correlating with the stage of oestrous reported in the vagina.  This finding may be related to the test article related bodyweight effects seen in females of this dose group during treatment, but was considered to be of uncertain relationship to treatment since the oestrus cycles prior to pairing, mating performance and fertility of these females had been unaffected by treatment. Overall the reproductive endpoints assessed which were unaffected by treatment included: oestrous cycles, pre-coital interval, mating performance, fertility, gestation length, offspring clinical condition, litter size, survival, sex ratio or external development. The NOAEL for fertility was determined to be >=15000 ppm, representing mean achieved doses of >=868.7 mg/kg bw/day.

Developmental toxicity: Survival, litter size and sex ratio were not adversely affected by parental exposure to 2,6-imethyloct 7-en-2-ol. There was no difference in offspring body weight gain from Day 1-4 of age when compared with Controls; growth thereafter (Day 4-7) was lower in male and female offspring from parents receiving 15000 ppm. This low growth of the offspring was considered not to represent an adverse effect of treatment since the survival, clinical condition and macropathology findings for the offspring were not affected by reatment. There were no changes detected at examination of the offspring dying before or at scheduled termination which indication any effect of the test material.

The study indicates that there were no developmental adverse effects attributed to treatment with 2,6-dimethyloct-7-en-2-ol (dihydromyrcenol) by dietary administration. The NOAEL for developmental toxicity was therefore determined to be the highest dose tested of >=15000 ppm, representing a mean achieved dose of 975.8 mg/kg bw/day via dams during gestation.

Reference 2

Endpoint:

screening for reproductive / developmental toxicity

Type of information:

read-across from supporting substance (structural analogue or surrogate)

Adequacy of study:

key study

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: Information is derived from an analogue

Justification for type of information:

Fertility effects of Tetrahydromyrcenol are derived from Dihydromyrcenol, a close analogue. The read across justification is presented in the Toxicity for reproduction Endpoint summary, the accompanying files are also attached there.

Reason / purpose for cross-reference:

read-across source

Key result

Dose descriptor:

NOAEL

Effect level:

>= 868.7 mg/kg bw/day (actual dose received)

Based on:

test mat.

Sex:

male

Basis for effect level:

other: NOAEL = Highest dose tested

Key result

Dose descriptor:

NOAEL

Remarks:

pre-mating

Effect level:

>= 954.5 mg/kg bw/day (actual dose received)

Based on:

test mat.

Sex:

female

Basis for effect level:

other: NOAEL = highest dose tested

Dose descriptor:

NOAEL

Remarks:

gestation

Effect level:

>= 975.8 mg/kg bw/day (actual dose received)

Based on:

test mat.

Sex:

female

Basis for effect level:

other: NOAEL = Highest dose tested

Dose descriptor:

NOAEL

Remarks:

lactation

Effect level:

>= 1 449.4 mg/kg bw/day (actual dose received)

Based on:

test mat.

Sex:

female

Basis for effect level:

other: NOAEL = Highest dose tested

Key result

Critical effects observed:

no

Key result

Dose descriptor:

NOAEL

Remarks:

developmental (gestation)

Generation:

F1

Effect level:

>= 975.8 mg/kg bw/day

Based on:

test mat.

Sex:

female

Basis for effect level:

other: NOAEL = Maximum tested dose level

Dose descriptor:

NOAEL

Remarks:

developmental (lactation)

Generation:

F1

Effect level:

>= 1 449.4 mg/kg bw/day

Based on:

test mat.

Sex:

female

Basis for effect level:

other: NOAEL = Maximum tested dose level

Key result

Reproductive effects observed:

no

Conclusions:

Under the conditions of this study, there are no adverse effects for fertility up the maximum dose: NOAEL for fertility was determined to be >=15000 ppm, representing mean achieved doses of 868.7 mg/kg/day.

Effect on fertility: via oral route

Endpoint conclusion:

no adverse effect observed

Study duration:

subacute

Species:

rat

Quality of whole database:

Reliable without restriction, guideline study

Additional information

The reproductive toxicity of Tetrahydromyrcenol is derived by read across from Dihydromyrcenol. First the full OECD TG 422 of Dihydromyrcenol is summarised. Thereafter the read across justification will be presented. The accompanying files are also attached here.

Dihydromyrcenol and its effects in a OECD TG 422

Dihydyromyrcenol is tested in an OECD TG 422 test. The repeated dose toxicity is repeated here and includes the developmental effects to present a complete picture. In the present executive summary the reproductive/developmental toxicity effects are reported following dietary administration. A similarly constituted control group was assigned, and received the vehicle, powdered SDS VRF1 certified diet with corn oil, throughout the same relative treatment period.

Reproductive females were treated daily for three weeks before pairing, throughout pairing, gestation and until Day 6 of lactation. Females were allowed to litter and rear their offspring to weaning and were killed on Day 7 of lactation. The F1 generation were killed on Day 7 of age, but did not directly receive the test substance; any exposure was in utero or via the milk. Toxicity phase males were treated daily for three weeks before pairing up to necropsy after a minimum of five consecutive weeks.

During the study, clinical condition, detailed physical examination and arena observations, sensory reactivity, grip strength, motor activity, body weight, food consumption, ophthalmic examination, haematology (peripheral blood), blood chemistry, urinalysis, oestrous cycles, pre-coital interval, mating performance, fertility, gestation length, organ weight and macroscopic pathology and histopathology investigations were undertaken. The clinical condition, litter size and survival, sex ratio,bodyweightand macropathology for all offspring were also assessed.

Results

Clinical signs: There were no adverse effects observed attributed to treatment on: clinical condition, sensory reaction, grip strength, motor activity, ophthalmic examination and macroscopic pathology.  

Other effects: Minor treatment related effects were observed for body weight, histopathology, blood chemistry and urine analysis. However, the animals showed recovery and, the survival, clinical condition and reproductive performance of the animals was unaffected by treatment, therefore they are considered not to represent an adverse effect of the treatment. In the kidney of male rats, alpha 2u-globulin nephropathy was observed. This is a male-rat-specific finding associated with the binding of xenobiotics to this protein and interference with the normal lysosomal uptake and degradation of the protein, and did not adversely affect the reproductive performance of the animals. Some histological changes were observed in the liver (portal pigment), also these effect were not considered adverse, as no effect on the survival or reproductive performance of the animals was observed and no indication of impaired liver function could be seen from the blood chemistry analyses.

Fertility effects: In the female reproductive tract, all females receiving 15000 ppm were observed to be in diestrus in comparison to the Controls and low and intermediate dosage groups. This was associated with a reduced incidence of luminal dilatation of the uterus, correlating with the stage of oestrous reported in the vagina.  This finding may be related to the test article related bodyweight effects seen in females of this dose group during treatment, but was considered to be of uncertain relationship to treatment since the oestrus cycles prior to pairing, mating performance and fertility of these females had been unaffected by treatment. Overall the reproductive endpoints assessed which were unaffected by treatment included: oestrous cycles, pre-coital interval, mating performance, fertility, gestation length, offspring clinical condition, litter size, survival, sex ratio or external development. The NOAEL for fertility was determined to be >=15000 ppm, representing mean achieved doses of >=868.7 mg/kg bw/day.

Developmental toxicity: Survival, litter size and sex ratio were not adversely affected by parental exposure to 2,6-imethyloct 7-en-2-ol. There was no difference in offspring body weight gain from Day 1-4 of age when compared with Controls; growth thereafter (Day 4-7) was lower in male and female offspring from parents receiving 15000 ppm. This low growth of the offspring was considered not to represent an adverse effect of treatment since the survival, clinical condition and macropathology findings for the offspring were not affected by treatment. There were no changes detected at examination of the offspring dying before or at scheduled termination which indication any effect of the test material.

The study indicates that there were no developmental adverse effects attributed to treatment with 2,6-dimethyloct-7-en-2-ol (dihydromyrcenol) by dietary administration. The NOAEL for developmental toxicity was therefore determined to be the highest dose tested of >=15000 ppm, representing a mean achieved dose of 975.8 mg/kg bw/day via dams during gestation.

The reproductive toxicity of Tetrahydromyrcenol (18479-57-7) using read across from Dihydromyrcenol 18479-58-8)

Introduction and hypothesis for the analogue approach

Tetrahydromyrcenol is a branched C8 alkyl chain to which a tertiary alcohol is attached at the end of the alkyl chain. For this substance no reproductive toxicity data are available. In accordance with Article 13 of REACH, lacking information should be generated whenever possible by means other than vertebrate animal tests, i. e. applying alternative methods such as in vitro tests, QSARs, grouping and read-across. For assessing the reproductive toxicity of Tetrahydromyrcenol the analogue approach is selected because for one closely related analogue reproductive toxicity information is available which can be used for read across.

Hypothesis: Tetrahydromyrcenol has the same reproductive toxicity as Dihydromyrcenol resulting in a similar NOAEL for fertility and/or developmental toxicity.

Available information: The source chemical Dihydromyrcenol has been tested in a well conducted dietary repeated dose toxicity test (OECD TG 422 under GLP, kl 1), resulting in a NOAEL of >= 868 mg/kg bw (maximum dietary concentration was 15000 ppm).

Target chemical and source chemical(s)

Chemical structures of the Tetrahydromyrcenol (the target) and are shown in the Data matrix including physico-chemical properties and toxicological information, thought relevant for reproductive toxicity.

Purity / Impurities

The purity and impurities of the target chemical do not indicate skin sensitisation potential other than indicated by the parent substance. The impurities are all below 10%.

Analogue approach justification

According to Annex XI 1.5 read across can be used to replace testing when the similarity can be based on a common backbone and a common functional group. When using read across the result derived should be applicable for C&L and/or risk assessment and it should be presented with adequate and reliable documentation.

Analogue selection: Dihydromyrcenol was the closest analogue for which reproductive toxicity information was available.

Structural similarities and differences: Tetrahydro and Dihydro-myrcenol both have the same2,6-dimethyl)octan-2-ol backbone, with a tertiary alcohol functional group. The difference is that Dihydromyrcenol has at one end a double (unsaturated) bond, while Tetrahydromyrcenol is fully saturated.

Toxico-kinetics: Oral absorption of both substances will be alike based on the similarity in chemical structure, molecular weight, being liquids and physico-chemical properties.Metabolism: The methyl groups of both substances may be oxidized into primary alcohols and acids. Reduction of the tertiary alcohols resulting in a double bond at this site is not too likely because conjugation at this site would become limited. Both tertiary and primary alcohols can be conjugated. The acidic metabolites may also be excreted unchanged (Belsito et al. (2010b). The double bond solely present in Dihydromyrcenol may via epoxidation turn into an alcohol are acid.

Toxico-dynamic: The chemicals structure, the functional groups and similarities of the metabolite indicate a similar mode of action for both fertility and developmental toxicity endpoints. Receptor binding may be important for reproductive endpoints. These are expected to be similar based on the similarities in structure as presented above.

Other experimental information to indicate the feasibility of the analogue: Tetrahydro and Dihydro-myrcenol are both negative in the Ames test, indicating absence of genotoxic features that may influence developmental toxicity. 

Remaining uncertainties:There are no remaining uncertainties as presented above.

Data matrix

The relevant information on physico-chemical properties and (reproductive) toxicological characteristics are presented in the Data Matrix in Table 1.

Conclusions for reproductive toxicity 

For Tetrahydromyrcenol the Reproductive toxicity can be derived from Dihydromyrcenol for which a dietary OECD TG 422, K1 is available). The derived NOAEL is >=868 mg/kg bw (15000 ppm) in absence of any fertility and developmental toxicity.

Final conclusion on fertility and developmental toxicity: For Tetrahydromyrcenol no fertility and no developmental toxicity is anticipated >= 868 mg/kg bw (>=15000 ppm) the limit dose. This means that ‘no adverse effects’ are observed. This value will be brought forward to the risk assessment.

References

D. Belsito, D. Bickers, M. Bruze, P. Calow, H. Greim, J.M. Hanifin, A.E. Rogers, J.H. Saurat, I.G. Sipes, H. Tagami: The RIFM Expert Panel. A safety assessment of branched chain saturated alcohols when usedas fragrance ingredients. Food and Chemical Toxicology 48 (2010) S1–S46

Data matrix: Information from Tetrahydromyrcenol and its analogue Dihydromyrcenol to supportthe read across

Common names	Tetrahydromyrcenol	Dihydromyrcenol
	Target	Source
Chemical structures
CAS no	18479-57-7	18479-58-8
REACH registration	For 2018	2010 registered.
Einecs	242-361-9	242-362-4
Molecular weight	158.29	156.27
Physico-chemical data
Physical state	Liquid	Liquid
Melting point, oC	<-20	<-20 (ECHA dissemination site)
Boiling point, oC	201.9	193(ECHA dissemination site)
Vapour pressure, Pa	9.3	20(ECHA dissemination site)
Water solubility, mg/l	281.9	939(ECHA dissemination site)
Log Kow	3.2 (m)	3.25(ECHA dissemination site)
Human health endpoints
Genotoxicity – Ames test	Not mutagenic (OECD TG 471)	Non mutagenic (OECD TG 471)
Reproductive toxicity, fertility inmg/kg bw	Read across from Dihydromyrcenol	NOAEL >=868 (OECD422)

 

Effects on developmental toxicity

Description of key information

Toxicity to development (OECDTG422): NOAEL >= 975.8 mg/kg bw/day (exposure via dams during gestation) (read-across)

Link to relevant study records

Reference

Reference 1

Endpoint:

developmental toxicity

Type of information:

experimental study

Adequacy of study:

key study

Study period:

25 March 2015 - 27 November 2015

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Justification for type of information:

This information is used for read across to Tetrahydromyrcenol.

Qualifier:

according to guideline

Guideline:

other: OECD Guideline 422 (Combined Repeated Dose Toxicity Study with the Reproduction / Developmental Toxicity Screening Test)

Version / remarks:

22 March 1996

GLP compliance:

yes (incl. QA statement)

Limit test:

no

Species:

rat

Strain:

other: Crl:CD(SD)

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Charles River (UK) Ltd.
- Females nulliparous and non-pregnant: Yes
- Age at study initiation: Approx. 10 weeks
- Weight at study initiation: Males: 335-402 g / Females: 230-291 g
- Fasting period before study: Only fasted overnight before blood sampling for haematology and blood
chemistry investigations
- Housing: Polycarbonate body with a stainless steel mesh lid (changed at appropriate intervals) used
during the acclimatisation, pre-pairing, gestation, littering and lactation periods. Polypropylene cages
used during pairing.
- Diet: Ad libitum (but removed overnight before blood sampling for haematology and blood chemistry
investigations)
- Water (e.g. ad libitum): Ad libitum (but removed overnight during urine collection)
- Acclimation period: Six days before start of treatment

DETAILS OF FOOD AND WATER QUALITY: SDS VRF1 Certified powdered diet; Potable water from
the public supply via polycarbonate bottles with sipper tubes. Bottles were changed at appropriate
intervals.

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 19-23
- Humidity (%): 40-70
- Air supply: Filtered fresh air which was passed to atmosphere and not recirculated
- Photoperiod (hrs dark / hrs light): 12/12

IN-LIFE DATES:
Animal arrival: 24 June 2015
F0 necropsy: Toxicity phase 13 August 2015, Recovery phase 28 August 2015, Reproductive phase
19 to 24 August 2015

Route of administration:

oral: feed

Vehicle:

corn oil

Remarks:

used as stabiliser

Details on exposure:

DIET PREPARATION
- Rate of preparation of diet (frequency): Weekly
- Mixing appropriate amounts with (Type of food): SDS VRF1 certified powdered diet
- Storage temperature of food: Ambient temperature (nominally 21°C) for eight days. Deep-frozen (nominally -20°C) for 22 days.

VEHICLE
- Justification for use and choice of vehicle (if other than water): The dietary of administration was chosen to simulate a condition of potential human exposure.
- Concentration in vehicle: test material to corn oil ratio 5:1

Analytical verification of doses or concentrations:

yes

Details on analytical verification of doses or concentrations:

For Week 1 and Final Week of treatment, representative samples of test diet were taken and submitted for analysis using chromatography. Each diet sample was sub-sampled (10 g ± 0.5 g) in duplicate and analysed in accordance with the analytical procedure. The analysed concentrations of 2,6-dimethyloct-7-en-2-ol in the test formulations analysed during the study, associated procedural recovery data and the deviation of mean results from nominal values were between -6% and +3.3% which is within applied limits of +10%/-15%. This confirms the accuracy of formulation. Procedural recovery values remained within 99.2% and 100.7%, confirming the continued accuracy of the method. The difference of individual values from the mean was <5% confirming the precision of analysis.

Details on mating procedure:

- M/F ratio per cage: 1:1 from within the same treatment groups.
- Length of cohabitation: Up to two weeks.
- Proof of pregnancy: Ejected copulation plugs in cage tray or sperm in the vaginal smear referred to as Day 0 of gestation
- After successful mating each pregnant female was caged: individually

Duration of treatment / exposure:

Reproductive phase females: Three weeks before pairing, then throughout pairing and gestation until Day 6 of lactation.
Toxicity phase males: Three weeks pre-pairing up to necropsy after minimum of five weeks.

Frequency of treatment:

Continuous

Duration of test:

6 weeks (5 weeks pre-mating / mating / gestation and 1 week postnatal)

Dose / conc.:

15 000 ppm

Remarks:

Corresponding actual intake
Week 1 to 3 prior to pairing: 954.5 mg/kg bw/day
Days 0-19 of gestation: 975.8 mg/kg bw/day
Days 1-6 of lactation: 1449.4 mg/kg bw/day
Males: 868.7 mg/kg bw/day

Dose / conc.:

5 000 ppm

Remarks:

Corresponding actual intake
Week 1 to 3 prior to pairing: 349.8 mg/kg bw/day
Days 0-19 of gestation: 342.7 mg/kg bw/day
Days 1-6 of lactation: 599.2 mg/kg bw/day
Males: 291.7 mg/kg bw/day

Dose / conc.:

1 500 ppm

Remarks:

Corresponding actual intake
Week 1 to 3 prior to pairing: 108 mg/kg/day
Days 0-19 of gestation: 102.3 mg/kg/day
Days 1-6 of lactation: 202.4 mg/kg/day
Males: 89 mg/kg bw/day

No. of animals per sex per dose:

10 females per dose group
5 males per dose group

Control animals:

yes, concurrent vehicle

yes, historical

Details on study design:

- Dose selection rationale: The dietary levels of 1500, 5000 and 15000 ppm were selected based on the results of a 21-day preliminary study (Envigo Study Number: CIZ0001). In that study, administration of 2,6-dimethyloct-7-en-2-ol to CD rats by dietary administration for 21 days at dietary levels of 1500, 7500 or 15000 ppm was well tolerated. No animals died and the clinical condition of the animals was satisfactory.
- Rationale for animal assignment: random (Note: On Day 1 of study all animals were weighed and body weights were reviewed before dosing commenced by Study Management. The groups were adjusted to reduce inter-/intra-group variation.)

Maternal examinations:

CAGE SIDE OBSERVATIONS: Yes
- Time schedule: Twice Daily

DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: Daily

BODY WEIGHT: Yes
Reproductive phase females:
- Before feeding on the day that treatment commenced, (Week 0) and weekly before pairing, Days 0, 6, 13 and 20 after mating, Day 1, 4, and 7 of lactation, On the day of necropsy.

OTHER EXAMINATIONS: During the study, clinical condition, detailed physical examination and arena observations, sensory reactivity, grip strength, motor activity, body weight, food consumpt ion, ophthalmic examination, haematology (peripheral blood), blood chemistry, urinalysis, oestrous cycles, pre-coital interval, mating performance, fertility, gestation length, organ weight and macroscopic pathology and histopathology investigations were undertaken.

Ovaries and uterine content:

Not studied

Fetal examinations:

Where possible, a fresh macroscopic examination (external and internal) with an assessment of stomach for milk content was performed.

Statistics:

Not relevant

Indices:

- Post-implantation survival index (%) = Total number of offspring born / Total number of uterine implantation sites x 100
- Live birth index (%) = Number of live offspring on Day 1 after littering / Total number of offspring born x 100
- Viability index (%) = Number of live offspring on Day 7 / Number of liver offspring on Day 1 after littering x 100

Historical control data:

Not reported for offspring

Clinical signs:

no effects observed

Mortality:

no mortality observed

Body weight and weight changes:

effects observed, treatment-related

Description (incidence and severity):

Reproductive phase Females: body weight gain was significantly reduced from Day 13-20 of gestation in females treated at 5000 or 15000 ppm.
Lactation phase Females: Day 1-7: body weight gain was significantly reduced in females treated at 15000 ppm.

Food consumption and compound intake (if feeding study):

effects observed, treatment-related

Description (incidence and severity):

During gestation, food intake in females treated at 15000 ppm food intake remained slightly low when compared with controls, from Day 0 to Day 3; thereafter food intake improved and was similar to that of controls. During lactation, food intake was slightly low in females treated at 5000 ppm with a markedly low food intake observed in females treated at 15000 ppm. This effect persisted throughout lactation (until Day 7).

Food efficiency:

not specified

Description (incidence and severity):

On occasions throughout the treatment period, animals treated at 15000 ppm were observed to consume more water than the animals treated at 0, 1500 or 5000 ppm. This effect is considered to be due to the palatability of the test material and not an adverse response to treatment.

Ophthalmological findings:

no effects observed

Description (incidence and severity):

Ophthalmic examination revealed no findings that could be attributed to treatment with 2,6- dimethyloct-7-en-2-ol.

Haematological findings:

effects observed, treatment-related

Description (incidence and severity):

None of the findings were conclusively linked to any of the treatment related microscopic pathology changes, and since all of the affected parameters evaluated during Week 2 of recovery showed recovery and, the survival, clinical condition and reproductive performance of the animals was unaffected by treatment, they are considered not to represent an adverse effect of treatment.

Females 5000 or 15000 ppm:
- Mean cell haemoglobin concentration was significantly increased (p<0.05 in both groups)
- Mean cell volume was significantly reduced in females treated at 15000 ppm

Clinical biochemistry findings:

effects observed, treatment-related

Description (incidence and severity):

None of the findings were conclusively linked to any of the treatment related microscopic pathology changes, and since all of the affected parameters evaluated during Week 2 of recovery showed recovery and, the survival, clinical condition and reproductive performance of the animals was unaffected by treatment, they are considered not to represent an adverse effect of treatment.

Females 1500, 5000 or 15000 ppm:
- glucose concentration significantly reduced (p<0.05, p<0.01 and p<0.01 respectively)
- potassium concentration significantly reduced (5000 or 15000 ppm) (p<0.01 both)
- albumin/globulin ratio reduced (15000 ppm)
- cholesterol increased (15000 ppm)

Urinalysis findings:

effects observed, treatment-related

Description (incidence and severity):

None of the findings were conclusively linked to any of the treatment related microscopic pathology changes, and since all of the affected parameters evaluated during Week 2 of recovery showed recovery and, the survival, clinical condition and reproductive performance of the animals was unaffected by treatment, they are considered not to represent an adverse effect of treatment.

Females 15000 ppm:
- increase in urinary volume (not significant)
- significant increase in total potassium (p<0.05)
- significant increase in total chloride levels (p<0.01)

Behaviour (functional findings):

no effects observed

Immunological findings:

not specified

Organ weight findings including organ / body weight ratios:

effects observed, treatment-related

Description (incidence and severity):

Females (1500, 5000 and 15000 ppm):
- Uterus, cervix, oviducts: absolute and adjusted combined weight showed a dose-proportional (but not statistically significant) reduction in weight (95%, 81% and 66% of control, respectively)
- Kidney: marginally low absolute and adjusted kidney weight (15000 ppm)

Gross pathological findings:

no effects observed

Neuropathological findings:

no effects observed

Histopathological findings: non-neoplastic:

effects observed, treatment-related

Description (incidence and severity):

Changes related to treatment with 2,6-dimethyloct-7-en-2-ol were seen in the female reproductive tract. All females receiving 15000 ppm were in diestrus, whereas Control females and females receiving 1500 or 5000 ppm were more often in proestrus or metoestrus. Uterine luminal dilatation was not evident in any of the females receiving 15000 ppm. The apparent difference in stage of oestrus is of uncertain significance. Note: This finding may be related to the test article related bodyweight effects seen in females of this dose group during treatment, but was considered to be of uncertain relationship to treatment since the efect was reversible and the oestrus cycles prior to pairing, mating performance and fertility of these females had been unaffected by treatment.

Histopathological findings: neoplastic:

no effects observed

Other effects:

no effects observed

Number of abortions:

not specified

Pre- and post-implantation loss:

no effects observed

Total litter losses by resorption:

no effects observed

Early or late resorptions:

not specified

Dead fetuses:

no effects observed

Changes in pregnancy duration:

no effects observed

Description (incidence and severity):

Migrated Data from removed field(s)
Field "Effects on pregnancy duration" (Path: ENDPOINT_STUDY_RECORD.DevelopmentalToxicityTeratogenicity.ResultsAndDiscussion.ResultsMaternalAnimals.MaternalDevelopmentalToxicity.EffectsOnPregnancyDuration): no effects observed

Changes in number of pregnant:

no effects observed

Other effects:

not specified

Key result

Dose descriptor:

NOAEL

Remarks:

gestation

Effect level:

>= 975.8 mg/kg bw/day (actual dose received)

Based on:

test mat.

Basis for effect level:

other: NOAEL = highest dose tested

Dose descriptor:

NOAEL

Remarks:

lactation

Effect level:

>= 1 449.4 mg/kg bw/day (actual dose received)

Based on:

test mat.

Basis for effect level:

other: NOAEL = highest dose tested

Key result

Abnormalities:

no effects observed

Fetal body weight changes:

not examined

Description (incidence and severity):

Migrated Data from removed field(s)
Field "Fetal/pup body weight changes" (Path: ENDPOINT_STUDY_RECORD.DevelopmentalToxicityTeratogenicity.ResultsAndDiscussion.ResultsFetuses.FetalPupBodyWeightChanges): effects observed, treatment-related
Field "Description (incidence and severity)" (Path: ENDPOINT_STUDY_RECORD.DevelopmentalToxicityTeratogenicity.ResultsAndDiscussion.ResultsFetuses.DescriptionIncidenceAndSeverityFetalPupBodyWeightChanges): There was no difference in offspring body weight gain from Day 1-4 of age when compared with the control group. Growth thereafter (Day 4-7) was lower in male and female offspring from parents receiving 15000 ppm. This low growth of the offspring was considered not to represent an adverse effect of treatment since the survival, clinical condition and macropathology findings for the offspring were not affected by treatment. There was no effect of treatment in offspring from parents receiving 5000 or 1500 ppm.

Reduction in number of live offspring:

no effects observed

Changes in sex ratio:

no effects observed

Changes in litter size and weights:

no effects observed

Changes in postnatal survival:

effects observed, non-treatment-related

Description (incidence and severity):

Mortality was observed in litters of all test groups (including control), but this is not considered treatment related.

External malformations:

no effects observed

Skeletal malformations:

not specified

Visceral malformations:

not specified

Other effects:

no effects observed

Key result

Dose descriptor:

NOAEL

Remarks:

gestation

Effect level:

>= 975.8 mg/kg bw/day (actual dose received)

Based on:

test mat.

Remarks:

received via dams

Sex:

male/female

Basis for effect level:

other: NOAEL = highest dose tested

Dose descriptor:

NOAEL

Remarks:

lactation

Effect level:

>= 1 449.4 mg/kg bw/day (actual dose received)

Based on:

test mat.

Remarks:

received via dams

Sex:

male/female

Basis for effect level:

other: NOAEL = highest dose tested

Key result

Developmental effects observed:

no

Conclusions:

Dihydromyrcenol did not show adverse developmental effects in an OECD TG 422 up to the maximum dose of 15000 ppm (equivalent to 975.8 mg/kg bw/day).

Executive summary:

In this endpoint the findings of the reproductive/developmental toxicity screening are reported as part of an OECD TG 422 oral toxicity study in rats following dietary administration of Dihydromyrcenol. A similarly constituted control group was assigned, and received the vehicle, powdered SDS VRF1 certified diet with corn oil, throughout the same relative treatment period. Reproductive females were treated daily for three weeks before pairing, throughout pairing, gestation and until Day 6 of lactation. Females were allowed to litter and rear their offspring to weaning and were killed on Day 7 of lactation. The F1 generation were killed on Day 7 of age, but received no direct administration of the substance; any exposure was in utero or via the milk. Males were treated daily for three weeks before pairing. During the study, clinical condition, detailed physical examination and arena observations, sensory reactivity, grip strength, motor activity, body weight, food consumption, ophthalmic examination, haematology (peripheral blood), blood chemistry, urinalysis, oestrous cycles, pre-coital interval, mating performance, fertility, gestation length, organ weight and macroscopic pathology and histopathology investigations were undertaken. The clinical condition, litter size and survival, sex ratio,bodyweightand macropathology for all offspring were also assessed.

Results

There were no adverse maternal effects observed attributed to treatment on: clinical condition, sensory reaction, grip strength, motor activity, ophthalmic examination and macroscopic pathology.  In addition oestrous cycles, pre-coital interval, mating performance, fertility, gestation length, offspring clinical condition, litter size, survival, sex ratio or external development were unaffected. In reproductive phase females, food intake was low in females treated at 15000 ppm throughout pre-pairing until Day 3 of gestation; thereafter food intake improved throughout the gestation phase. However, body weight gain was significantly reduced from Day 13-20 of gestation in females treated at 5000 or 15000 ppm. For females treated at 15000 ppm, this effect persisted throughout lactation (Day 1-7) when food consumption was low, which was likely responsible for the low growth of the offspring from Day 4 to Day 7 of age. For females treated at 5000 ppm, body weight gain during lactation was similar to the control group despite food intake being slightly low.

Other effects: Several minor differences in haematology, blood chemistry and urinalysis parameters were recorded; none were conclusively linked to any of the treatment related microscopic pathology and since the survival, clinical condition and reproductive performance of the animals was unaffected by treatment, they are considered not to represent an adverse effect of treatment.

In the female reproductive tract, all females receiving 15000 ppm were observed to be in diestrus. This was associated with a reduced incidence of luminal dilatation of the uterus, correlating with the stage of oestrous reported in the vagina.  This finding may be related to the test article related bodyweight effects seen in females of this dose group during treatment, but was considered to be of uncertain relationship to treatment since the oestrus cycles prior to pairing, mating performance and fertility of these females had been unaffected by treatment.

Developmental toxicity: Survival, litter size and sex ratio were not adversely affected by parental exposure. There was no difference in offspring body weight gain from Day 1-4 of age when compared with Controls; growth thereafter (Day 4-7) was lower in male and female offspring from parents receiving 15000 ppm. This low growth of the offspring was considered not to represent an adverse effect of treatment since the survival, clinical condition and macropathology findings for the offspring were not affected by reatment. There were no changes detected at examination of the offspring dying before or at scheduled termination which indication any effect of the test material.

Conclusion: the study indicates that there were no developmental adverse effects attributed to treatment with the substance by dietary administration. The NOAEL for developmental toxicity was therefore determined to be the highest dose tested of 15000 ppm, representing a mean achieved dose of 975.8 mg/kg bw/day via dams during gestation.

Reference 2

Endpoint:

developmental toxicity

Type of information:

read-across from supporting substance (structural analogue or surrogate)

Adequacy of study:

key study

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: Information is derived from an analogue

Justification for type of information:

The information for Tetrahydromyrcenol is derived from Dihydromyrcenol. The read across justification is presented in the Toxicity to reproduction Endpoint summary. The accompanying files are also attached there.

Reason / purpose for cross-reference:

read-across source

Key result

Dose descriptor:

NOAEL

Remarks:

gestation

Effect level:

>= 975.8 mg/kg bw/day (actual dose received)

Based on:

test mat.

Basis for effect level:

other: NOAEL = highest dose tested

Key result

Dose descriptor:

NOAEL

Remarks:

lactation

Effect level:

>= 1 449.4 mg/kg bw/day (actual dose received)

Based on:

test mat.

Basis for effect level:

other: NOAEL = highest dose tested

Key result

Abnormalities:

no effects observed

Key result

Dose descriptor:

NOAEL

Effect level:

>= 975.8 mg/kg bw/day (actual dose received)

Based on:

test mat.

Remarks:

received via dams

Sex:

male/female

Basis for effect level:

other: NOAEL = highest dose tested

Key result

Dose descriptor:

NOAEL

Effect level:

>= 1 449.4 mg/kg bw/day (actual dose received)

Based on:

test mat.

Remarks:

received via dams

Sex:

male/female

Basis for effect level:

other: NOAEL = highest dose tested

Key result

Developmental effects observed:

no

Conclusions:

The NOAEL for developmental toxicity derived from Dihydromyrcenol was >=15000 ppm, representing a mean achieved dose of 975.8 mg/kg bw/day via dams during gestation.

Effect on developmental toxicity: via oral route

Endpoint conclusion:

no adverse effect observed

Study duration:

subacute

Species:

rat

Quality of whole database:

Reliable without restriction, guideline study

Additional information

In the fertility section above also the developmental toxicity is included.

Justification for classification or non-classification

Based on the available data, TetrahydroMyrcenol does not need to be classified for reproduction and developmental toxicity in accordance with the criteria outlined in EU CLP (1272/2008/EC and its updates).