Alcohols, C13-15-branched and linear

Endpoint summary

Key value for chemical safety assessment

Effects on fertility

Description of key information

OECD 422 (BASF, 2022): NOAEL = 1000 mg/kg bw for systemic toxicicty and reproductive performance.

There is currently no further data available for CAS 85566 -16 -1 or its structural analogues adressing this endpoint. However, there is a testing proposal for an OECD 443 guideline EOGRT study for the read-across substance CAS 27458 -92 -0 currently under evaluation. Together with the OECD 422 guideline screening study conducted in 2022 with CAS 85566 -16 -1, a future read-across will be feasible to adress this endpoint.

Link to relevant study records

Reference

Endpoint:

screening for reproductive / developmental toxicity

Type of information:

experimental study

Adequacy of study:

key study

Study period:

May 2021-Jun 2022

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Reason / purpose for cross-reference:

reference to same study

Qualifier:

according to guideline

Guideline:

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

Version / remarks:

29 Jul 2016

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Limit test:

yes

Specific details on test material used for the study:

The analyses of the test item (= test substance) have been carried out at Competence Center Analytics, BASF SE, 67056 Ludwigshafen, Germany.
Name of test substance: C13-C15-Alcohol
CAS No.: 85566-16-1
Test substance No.: 20/0140-2
Batch identification: TK2004-02122020
Purity: 100% UVCB
Identity: confirmed
Homogeneity: given (visually)
Storage stability: Expiry date: 02 Jun 2022

ADDITIONAL TEST SUBSTANCE INFORMATION
Chemical name: Alcohols, C13-C15-branched and linear
Physical state/appearance: liquid / colorless, clear
Storage conditions: Room temperature

Species:

rat

Strain:

Wistar

Details on species / strain selection:

The test guideline requires the rat to be used as the animal species. This rat strain was selected since extensive historical control data are available for Wistar rats.

Sex:

male/female

Details on test animals or test system and environmental conditions:

Male and female Wistar rats, strain Crl:WI(Han), supplied by Charles River Laboratories, Research Models and Services, Germany GmbH, which were free from any clinical signs of disease, were used for the investigations. The females were nulliparous and non-pregnant at the beginning of the study. The receipt of males (about 77-90 days old) and females (about 70-76 days old) at different age warrants that no sibling males and females will be paired during the study. These animals were used as F0 generation parental animals. All other animals used in this study (F1 generation pups) were derived from the supplier provided animals.

HOUSING AND DIET
During the pretreatment, premating, mating and postmating period of the study, the rats were housed together (up to 5 [pretreatment] and 2 animals [premating] per sex and cage and 2 animals [males only during mating and postmating] per cage, respectively) in Polysulfonate cages Typ 2000P (H-Temp) supplied by TECNIPLAST, Hohenpeißenberg, Germany.
During the mating, gestation and lactation period and after weaning, the female rats were housed individually in Polycarbonate cages type III supplied by TECNIPLAST, Hohenpeißenberg, Germany and Becker & Co., Castrop-Rauxel, Germany, with the following exceptions:
• During overnight matings, male and female mating partners were housed together in Polycarbonate cages type III.
• Pregnant animals and their litters were housed together until PND 13 in Polycarbonate cages type III.
Pregnant females were provided with nesting material (compacted fibers of softwood, Typ SAFE® compact nesting large, J. Rettenmaier & Söhne GmbH + Co KG, Rosenberg, Germany) toward the end of gestation.
For enrichment wooden gnawing blocks (Typ SAFE® block large, J. Rettenmaier & Söhne GmbH + Co KG, Rosenberg, Germany) were added. In addition, in Polysulfonate cages large play tunnels (Art. 14153, supplied by PLEXX B.V., Elst, The Netherlands) were added.
The cages with the test animals were arranged on the racks in such a way that uniform experimental conditions (ventilation and light) were ensured.
The animals were housed in fully air-conditioned rooms in which central air conditioning guaranteed a range of temperature of 20-24°C and a range of relative humidity of 45-65%. The air change rate was 15 times per hour. There were no or only minimal deviations from these limits.
The day/night cycle was 12 hours light from 06.00 h to 18.00 h and 12 hours darkness from 18.00 h to 06.00 h.
Before the study started, the animal room was completely disinfected using a disinfector (Geschko MLT 17 hydrogen peroxide gas generator (PEA; Germany)). The walls and the floor were cleaned once a week with water containing an appropriate disinfectant.
The food used was mouse and rat maintenance diet “GLP”, supplied by Granovit AG, Kaiseraugst, Switzerland, which was available to the animals ad libitum throughout the study (from the day of supply to the day before necropsy). Drinking water was supplied from water bottles (ad libitum).
Dust-free wooden bedding was used in this study (the present supplier is documented in the raw data).

Route of administration:

oral: gavage

Vehicle:

CMC (carboxymethyl cellulose)

Remarks:

0.5% CMC suspension in deionized water with 10 mg/100 mL Tween 80

Details on exposure:

The test substance suspensions in 0.5% CMC suspension in deionized water with 10 mg/100 mL Tween 80 were prepared in intervals, which took into account the analytical results of the stability verification.
For the preparation of the administration suspensions, the specific amount of test substance was weighed in a calibrated beaker, topped up with 0.5% CMC suspension in deionized water with 10 mg/100 mL Tween 80 and intensely mixed with the magnetic stirrer.
Before and during administration, the preparations were kept homogeneous with a magnetic stirrer.

Details on mating procedure:

Pairing of F0 generation parental animals
In general, each of the male and female animals was mated overnight in a 1:1 ratio for a maximum of 2 weeks. Throughout the mating period, each female animal was paired with a predetermined male animal from the same dose group.
The animals were paired by placing the male in the cage of the female mating partner from about 16:00 h until 06:30 - 09:00 h of the following morning. Deviations from the specified times were possible on weekends and public holidays and were reported in the raw data. A vaginal smear was prepared after each mating and examined for the presence of sperm. If sperm was detected, pairing of the animals was discontinued. The day on which sperm were detected was denoted " gestation day (GD) 0" and the following day "gestation day (GD) 1"

Standardization of litters (F1 generation pups)
On PND 4, the individual litters were standardized in such a way that, where possible, each litter contained 4 male and 4 female pups (always the first 4 pups/sex and litter were taken for further rearing). If individual litters did not have 4 pups/sex, the litters were processed in such a way that the most evenly distributed 8 pups per litter were present for further rearing (e.g., 5 male and 3 female pups). Surplus pups were sacrificed.
Standardization of litters was not performed in litters with =<8 pups.

Pups after standardization
On PND 4, as a result of standardization, the surplus pups were sacrificed under isoflurane anesthesia by decapitation. Blood was sampled for determination of thyroid hormone concentrations. All pups were examined externally and eviscerated; their organs were assessed macroscopically.

Analytical verification of doses or concentrations:

yes

Details on analytical verification of doses or concentrations:

The analyses were carried out at the Analytical Chemistry Laboratory of Experimental Toxicology and Ecology of BASF SE, Ludwigshafen, Germany.
Analytical verifications of the stability of the test substance in 0.5% CMC suspension in deionized water with 10 mg/100 mL Tween 80 over a period of 7 days at room temperature had been verified before the start of the study.
At the beginning (during premating), once during gestation and once during lactation of the study each 3 samples were taken from the lowest and highest concentration for potential homogeneity analyses. The 3 samples were withdrawn from the top, middle and bottom of the preparation vessel. These samples were used as a concentration control at the same time. At the above mentioned time points additionally one sample from the mid concentration was taken for concentration control analysis.
All test samples, plus a duplicate set of reserve samples, were withdrawn by staff of the Laboratory Reproduction Toxicology. The samples collected at the beginning of the administration period and during lactation were analyzed in the Analytical Laboratory.
The samples of the gestation were not analyzed because no relevant imprecision occurs during the analyses of the samples from the beginning and lactation of the study.
Reserve samples were stored at the Laboratory Reproduction Toxicology frozen (at -20°C).
Analyses of these samples were performed in case of equivocal analytical results with the original samples or after loss of/damage to original samples after agreement by the Study Director. Following finalization of the report, all analytical samples, including reserve samples, were discarded.

RESULTS
Stability analysis
The stability of test substance in 0.5% CMC suspension in deionized water (with 10 mg/100 mL Tween 80) was demonstrated for a period of 7 days at room temperature.

Homogeneity analysis
The homogeneous distribution of the test substance in 0.5% CMC suspension in deionized water (with 10 mg/100 mL Tween 80) was demonstrated.

Concentration control analysis
Measured values for C13-C15-Alcohol were in the expected range of the target concentrations (90–110 %) demonstrating the correctness of the preparations .

Frequency of treatment:

once daily

Details on study schedule:

The male and female rats were about 10 - 12 weeks old, when they arrived from the breeder. During an acclimatization period of about 3 weeks, estrous cycle determination prior to treatment was performed in a pool of up to 50 non-randomized female animals. Animals with no regular estrous cycle and/or animals with the lowest and highest body weights were eliminated from the study and used for other purposes. The 40 male and 40 female animals included in the study were about 13 - 15 weeks old at the beginning of treatment and their weight variation did not exceed 20 percent of the mean weight of each sex.
The assignment of the animals to the different test groups was carried out using a randomization program, according to their weight one day before the beginning of the administration period (day -1).
After the acclimatization period, the test substance was administered to the parental animals orally by gavage, once daily at approximately the same time in the morning. Females in labor were not treated. The treatment lasted up to one day prior to sacrifice. The animals of the control group were treated with the vehicle (0.5% CMC suspension in deionized water with 10 mg/100 mL Tween 80), in the same way. The volume administered each day was 10 mL/kg body weight. The calculation of the administration volume was based on the most recent individual body weight.
A detailed clinical observation (DCO) was performed in all animals once before the start of the administration period and thereafter at weekly intervals.
Males and females from the same dose group were mated, after two weeks of treatment, overnight at a ratio of 1:1.
The females were allowed to deliver and rear their pups until PND 4 (standardization) or PND 13.
Blood samples were taken from all surplus pups at PND 4 as well as from one male and one female pup per litter at PND 13 by decapitation under isoflurane anesthesia for hormone measurement.
On study day 30, a functional observational battery and motor activity measurement were carried out in five male animals per group.
On study day 55, a functional observational battery and motor activity measurement were carried out in five female animals (with litter) per group.
Blood samples were taken from all F0 parental male animals of all test groups shortly before sacrifice and from all F0 parental female animals of all test groups on PND 14.
The male and female animals were sacrificed 36 and 57 days, respectively, after the beginning of the administration, and examined.

Dose / conc.:

100 mg/kg bw/day (nominal)

Remarks:

Dose Group 1

Dose / conc.:

300 mg/kg bw/day (nominal)

Remarks:

Dose Group 2

Dose / conc.:

1 000 mg/kg bw/day (nominal)

Remarks:

Dose Group 3

No. of animals per sex per dose:

10

Control animals:

yes, concurrent vehicle

Parental animals: Observations and examinations:

Mortality
A check for moribund or dead animals was made twice daily on working days or once daily (Saturday, Sunday or on public holidays). If animals were in a moribund state, they were sacrificed and necropsied. The examinations of these animals were carried out according to the methods established at the BASF SE Laboratory for Pathology, Experimental Toxicology and Ecology, Ludwigshafen, Germany.

Clinical observations
A cageside examination was conducted at least daily for any signs of morbidity, pertinent behavioral changes and signs of overt toxicity before the administration as well as within 2 hours and between 2 and 5 hours after the administration. Abnormalities and changes were documented daily for each animal.
The parturition and lactation behavior of the dams was generally evaluated in the mornings in combination with the daily clinical inspection of the dams. Only particular findings (e.g. inability to deliver) were documented on an individual dam basis.
On weekdays (except Saturday, Sunday and public holidays) the parturition behavior of the dams was inspected in the afternoons in addition to the evaluations in the mornings.
The day of parturition was considered the 24-hour period from about 15:00 h of one day until about 15:00 h of the following day.

Food consumption
Generally, food consumption was determined once a week for male and female parental animals, with the following exceptions:
• Food consumption was not determined after the 2nd premating week (male parental animals).
• Food consumption of the females with evidence of sperm was determined on GD 0-7, 7-14 and 14-20.
• Food consumption of the females which gave birth to a litter was determined on PND 1-4, 4-7, 7-10 and 10-13.
Food consumption was not determined in females without positive evidence of sperm during the mating and the gestation period and in females without litter during the lactation period.

Body weight data
In general, the body weight of the male and female parental animals was determined once a week at the same time of the day (in the morning) until sacrifice, with the following exceptions:
• During the mating period the parental females were weighed on the day of positive evidence of sperm (GD 0) and on GD 7, 14 and 20.
• Females with litter were weighed on the day of parturition (PND 0) and on PND 1, 4, 7, 10 and 13.
The body weight change of the animals was calculated from these results.
Females without positive evidence of sperm, without litter and females after weaning (PND 13) were weighed weekly. These body weight data were solely used for the calculations of the dose volume.

Detailed clinical observations
Detailed clinical observations (DCO) were performed in all animals once prior to the first administration (day 0) and at weekly intervals during the administration period. The examinations started in the morning. The findings were ranked according to the degree of severity, if applicable. For observation, the animals were removed from their cages by the investigator and placed in a standard arena (50 × 37.5 × 25 cm). The following parameters listed were assessed:
1. Abnormal behavior in “handling”
2. Fur
3. Skin
4. Posture
5. Salivation
6. Respiration
7. Activity/arousal level
8. Tremors
9. Convulsions
10. Abnormal movements
11. Gait abnormalities
12. Lacrimation
13. Palpebral closure
14. Exophthalmos (Protruding eyeball)
15. Assessment of the feces excreted during the examination (appearance/consistency)
16. Assessment of the urine excreted during the examination
17. Pupil size

Functional observation battery
A functional observational battery (FOB) was performed in first surviving 5 male and selected surviving 5 female animals with litter per group at the end of the administration period starting at about 10:00 h. The FOB started in a randomized sequence with passive observations without disturbing the animals, followed by removal from the home cage, open field observations in a standard arena and sensorimotor tests as well as reflex tests. The findings were ranked according to the degree of severity, if applicable. The observations were performed at random.

Home cage observations
The animals were observed in their closed home cages (for a short period: about 10-30 seconds); any disturbing activities (touching the cage or rack, noise) were avoided during these examinations in order not to influence the behavior of the animals. Attention was paid to:
1. Posture
2. Tremors
3. Convulsions
4. Abnormal movements
5. Gait
6. Other findings

Open field observations
The animals were transferred to a standard arena (50 × 50 × 25 cm) and observed. The following parameters were examined:
1. Behavior on removal from cage
2. Fur
3. Skin
4. Salivation
5. Nasal discharge
6. Lacrimation
7. Eyes/pupil size
8. Posture
9. Palpebral closure
10. Respiration
11. Tremors
12. Convulsions
13. Abnormal movements/stereotypy
14. Gait
15. Activity/arousal level
16. Feces (appearance/ consistency) within 2 minutes
17. Urine (amount/color) within 2 minutes
18. Rearing within 2 minutes
19. Other findings

Sensory motor tests/Reflexes
The animals were removed from the open field and subjected to following sensory motor or reflex tests:
1. Reaction to an object being moved towards the face (Approach response)
2. Touch sensitivity (Touch response)
3. Vision (Visual placing response)
4. Pupillary reflex
5. Pinna reflex
6. Audition (Startle response)
7. Coordination of movements (Righting response)
8. Behavior during handling
9. Vocalization
10. Pain perception (Tail pinch)
11. Other findings
12. Grip strength of forelimbs
13. Grip strength of hindlimbs
14. Landing foot-splay test

Motor activity measurement
The measurement of motor activity (MA) was measured at the end of the administration period in first surviving 5 male and selected surviving 5 female animals with litter per group. Motor activity (MA) was measured from 14:00 h onwards on the same day as the FOB was performed. The examinations were performed using the TSE Labmaster System supplied by TSE Systems GmbH, Bad Homburg, Germany. For this purpose, the animals were placed in new clean polycarbonate cages with a small amount of bedding for the duration of the measurement. Eighteen beams were allocated per cage. The number of beam interrupts were counted over 12 intervals for 5 minutes per interval. The sequence in which the animals were placed in the cages was selected at random. On account of the time needed to place the animals in the cages, the starting time was "staggered" for each animal. The measurement period began when the 1st beam was interrupted and was finished exactly 1 hour later. No food or water was offered to the animals during these measurements and the measurement room was darkened after the transfer of the last animal.

Oestrous cyclicity (parental animals):

For all females in a pool of up to 50 animals, estrous cycle normality was evaluated before the randomization.
For a minimum of 2 weeks prior to mating estrous cycle length was evaluated by daily analysis of vaginal smear for all F0 female parental rats. Determination was continued throughout the pairing period until the female exhibited evidence of copulation.
At necropsy, an additional vaginal smear was examined to determine the stage of estrous cycle for each F0 female with scheduled sacrifice.

Sperm parameters (parental animals):

The pairing partners, the number of mating days until vaginal sperm was detected in the female animals, and the gestational status of the females were recorded for F0 breeding pairs.
For females the pairing partners, the number of mating days until vaginal sperm was detected and gestational status was recorded for F0 females.

Litter observations:

Pup clinical observations
The live pups were examined daily for clinical symptoms (including gross-morphological findings) during the clinical inspection of the dams and documented for each pup.

Pup body weight data
The pups were weighed on the day after birth (PND 1) and on PND 4 (before standardization), 7 and 13. Pups' body weight change was calculated from these results.
The individual weights were always determined at about the same time of the day (in the morning) and on PND 4 immediately before standardization of the litters.
“Runts” were defined on the basis of the body weights on PND 1. "Runts" are pups that weigh less than 75% of the mean weight of the respective control pups.

Nipple/areola anlagen
All surviving male pups were examined for the presence of nipple/areola anlagen on PND 13. The number of nipple/areola anlagen was counted.

Postmortem examinations (parental animals):

See any other information below

Postmortem examinations (offspring):

Pup necropsy observations
On PND 4, as a result of standardization, the surplus pups were sacrificed under isoflurane anesthesia by decapitation. Blood was sampled for determination of thyroid hormone concentrations. After sacrifice, the pups were examined externally and eviscerated, and the organs were assessed macroscopically.
On PND 13, one selected male and one female pup per litter was sacrificed under isoflurane anesthesia by decapitation. Blood was sampled for determination of thyroid hormone concentrations. Thyroid glands/parathyroid glands were fixed in neutral buffered 4% formaldehyde solution and were transferred to the Pathology Laboratory for possible further processing. The remaining bodies were necropsied as far as possible.
The remaining pups were sacrificed under isoflurane with CO2. After sacrifice, all pups were examined externally and eviscerated, and their organs were assessed macroscopically.
All stillborn pups and all pups that died before weaning were examined externally, eviscerated and their organs were assessed macroscopically.
All pups without notable findings or abnormalities were discarded after their macroscopic evaluation. Animals with notable findings or abnormalities were evaluated on a case-by-case basis, depending on the type of finding noted.

Statistics:

Food consumption (parental animals), body weight and body weight change (parental animals and pups; for the pup weights, the litter means were used), gestation days, anogenital distance, anogenital index: Simultaneous comparison of all dose groups with the control group using the DUNNETT-test (twosided) for the hypothesis of equal means.
Male and female mating indices, male and female fertility indices, females mated, females delivering, gestation index (females with liveborn pups), females with stillborn pups, females with all stillborn pups: Pair-wise comparison of each dose group with the control group using FISHER'S EXACT test (one-sided) for the hypothesis of equal proportions.
Mating days until day 0 pc, %postimplantation loss, pups stillborn, %perinatal loss, nipple development: Pair-wise comparison of the dose group with the control group using the WILCOXON test (one-sided+) with BONFERRONI-HOLM adjustment for the hypothesis of equal medians.
Implantation sites, pups delivered, pups liveborn, live pups day x, viability Index, survival index: Pair-wise comparison of the dose group with the control group using the WILCOXON test (one-sided-) with BONFERRONI-HOLM adjustment for the hypothesis of equal medians.
% live male day x, %live female day x: Comparison of the dose group with the control group was performed using the WILCOXON test (two-sided) for the hypothesis of equal medians.
Number of cycles and Cycle Length, rearing, grip strength of forelimbs and hindlimbs, landing foot-splay test, motor activity: Non-parametric oneway analysis using KRUSKAL-WALLIS test (two-sided). If the resulting p-value was equal or less than 0.05, a pairwise comparison of each dose group with
the control group was performed using WILCOXON-test (twosided) for the equal medians.

Reproductive indices:

See any other information below

Offspring viability indices:

Pup number and status at delivery
All pups delivered from the F0 parents (F1 litter) were examined as soon as possible on the day of birth to determine the total number of pups, the sex and the number of liveborn and stillborn pups in each litter. At the same time, the pups were also being examined for macroscopically evident changes. Pups, which died before this initial examination, were defined as stillborn pups.

Pup viability/mortality
In general, a check was made for any dead or moribund pups twice daily on workdays (once in the morning and once in the afternoon) or as a rule, only in the morning on Saturdays, Sundays or public holidays.
The number and percentage of dead pups on the day of birth (PND 0) and of pups dying between PND 1-4, 5-7 and 8-13 were determined. Pups which died accidentally or had to be sacrificed due to maternal death were not included in these calculations. The number of live pups/litter was calculated on the day after birth (PND 0), and on lactation days 4, 7 and 13. Furthermore, viability and survival indices were calculated according to the following formulas:

Viability index (%) = (number of live pups on day 4* after birth / number of live pups on the day of birth) x 100
* before standardization of litters (i.e. before culling)

Survival index (%) = (number of live pups on day 13 after birth / number of live pups on day 4* after birth) x 100
* after standardization of litters (i.e. after culling)

Clinical signs:

effects observed, treatment-related

Description (incidence and severity):

All high- and mid-dose males (10 out of 10, respectively) and three (out of 10) low-dose males showed salivation after dosing during the entire study. Salivation occurred immediately after dosing (up to 2 hours post dosing), in seven high-dose males even afterwards (between 2 and 5 hours post dosing).
Salivation was also observed in all high-dose females during premating, gestation and lactation period and in six high-dose females during mating. In this test group, the finding occurred immediately after dosing and in nine (during gestation) or one (lactation) female(s) even afterwards (between 2 and 5 hours post dosing). Furthermore, in the mid-dose group, four females showed salivation during premating, five females during mating, nine females during gestation and all ten females during lactation period. The temporary salivation is considered to be treatment-related, most likely as a local irritation of the upper digestive tract or as a result of the bad taste of the test substance/vehicle preparation.
No other clinical signs or changes of general behavior, which may be attributed to the test substance, were detected in any of the male and female F0 parental animals in any of the test groups 1-3 (100, 300 and 1000 mg/kg bw/d) during the study.
There were some spontaneous findings recorded which were not associated with the test substance treatment. For one high-dose male (No. 37), piloerection was recorded on in-life day 25.
Low-dose female No. 118 showed a labored respiration (slight on GD 17, moderate on GD 18-19), piloerection (GD 17-23), hypothermia (GD 18-19) and a red encrusted nose (GD 19) during gestation as well as a reduced nutritional condition (moderate on PND 0, slight on PND 1) and piloerection (PND 0-2) during lactation period. This dam lost its litter completely on PND 1.
For mid-dose female No. 130, all pups were stillborn.

Detailed clinical observations
Apart from transient salivation in three low-dose, all mid- and all high-dose males, and in four mid-dose and all high-dose females, no clinical signs or changes of general behavior which may be attributed to the test substance were detected in any male and female animals in any of the test groups.
One high-dose male showed piloerection during the DCO, this was considered unrelated to treatment.

Dermal irritation (if dermal study):

not examined

Mortality:

no mortality observed

Description (incidence):

There were no test substance-related or spontaneous mortalities in any of the groups.

Body weight and weight changes:

effects observed, non-treatment-related

Description (incidence and severity):

In all test substance-treated groups, mean body weights of all male and all female parental animals as well as mean body weight change of all parental female animals were comparable to the concurrent control values during the entire study.
Mean body weight change of the high-dose parental males was statistically significantly below the concurrent control values during in-life days 0-7, 13-21 and 0-35. As the body weight values of this animals are comparable with the control values (maximum about -3% below control animals) this perceived finding was assessed as incidental and not toxicological relevant.
The slight, but statistically significant decrease of body weight change in the low-dose parental males during in-life days 13-21 was assessed as not treatment-related since it was not related to dose.

Food consumption and compound intake (if feeding study):

effects observed, non-treatment-related

Description (incidence and severity):

Mean food consumption of male animals in the high dose group was statistically significantly increased during in-life days 7-13 (about 43.7% above control).
In the high-dose female animals, mean food consumption was statistically significantly decreased on in-life days 0-7 (about -12.5% below control). Afterwards food consumption was comparable to the concurrent control throughout gestation and lactation periods.
Both isolated events (decreases/increases) were not related to the dose, they were considered to be spontaneous in nature and not treatment related.
Food consumption values of the low- and mid-dose males and females during the entire study were comparable to the concurrent control values.

Food efficiency:

not examined

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

not examined

Ophthalmological findings:

not examined

Haematological findings:

effects observed, non-treatment-related

Description (incidence and severity):

No treatment-related, adverse changes among hematological parameters were observed.
At PND14, in dams of test groups 2 and 3 (300 and 1000 mg/kg bw/d) absolute and relative eosinophil counts were significantly decreased. However, absolute eosinophil counts were not changed dose-dependently. Absolute as well as relative eosinophil counts were marginally below historical control ranges (dams PND14, absolute eosinophils 0.08-0.19 Giga/L, relative eosinophil counts 1.2-2.8 %). However, this was the only changed differential blood cell fraction. Therefore, it was regarded as treatment related, but non-adverse (ECETOC Technical Report No. 85, 2002).
Additionally, in females of test group 1 (100 mg/kg bw/d) absolute monocyte counts were significantly increased, and in males of this test group relative monocyte counts were significantly higher compared to controls. Both changes were not dose dependent. These mentioned alterations were regarded as incidental and not treatment related.

Clinical biochemistry findings:

effects observed, non-treatment-related

Description (incidence and severity):

No treatment-related, adverse changes among clinical chemistry parameters were observed.
At the end of the administration period in dams of test group 3 (1000 mg/kg bw/d) total protein and albumin values were significantly decreased. Total protein values were below the historical control range whereas albumin values were within this range (dams PND14, total protein 58.64-63.56 g/L, albumin 34.02-38.67 g/L). Globulins were not significantly changed among these individuals. Because total protein is the sum of albumin and globulin values, and albumin levels were within the historical control range, the finding was regarded as marginal and non-adverse if at all treatment related (ECETOC Technical Report No. 85, 2002).
In males of test group 3, urea values were significantly decreased, but the values were within the historical control range (males, urea 4.14-6.84 mmol/L). In dams of test groups 1 and 2 (100 and 300 mg/kg bw/d) glucose levels were significantly increased, and in males of test group 2 creatinine values were significantly higher compared to controls. However, both alterations were not dose dependent. Therefore, the clinical chemistry changes in this paragraph were regarded as incidental and not treatment related.

Endocrine findings:

no effects observed

Description (incidence and severity):

Thyroid hormones
In parental males as well as in male and female pups at PND13 (test groups 1, 2 and 3; 100, 300 and 1000 mg/kg bw/d) no treatment-related alterations of T4 and TSH levels were observed.

Urinalysis findings:

not examined

Behaviour (functional findings):

effects observed, non-treatment-related

Description (incidence and severity):

Functional observational battery

Home cage observations
No test substance-related or spontaneous findings were observed in male and female animals of all test groups during the home cage observation.

Open field observations
The open field observations did not reveal any test substance-related findings in male and female animals of all test groups.

Sensorimotor tests/reflexes
There were no test substance-related findings in male and female animals of all test groups.

Quantitative Parameters
No test substance-related impaired parameters were observed in male and female animals of all test groups.

Motor activity measurement
No treatment-related, adverse changes on motor activity data (summation of all intervals) was observed in the male and female animals of all test substance-treated groups in comparison to the concurrent control values.
The mean number of beam interrupts was statistically significantly below the concurrent control values during interval 8 in the high-dose males and during interval 5 in the mid-dose females. However, the mean number of beam interrupts was statistically significantly above the concurrent control during interval 12 in the low-dose males.
Since the summary values (interval 1-12) were comparable to the control values in males and females, respectively, and these isolated events (decreases/increases) were not related to the dose, they were considered to be spontaneous in nature and not treatment related.

Immunological findings:

not examined

Organ weight findings including organ / body weight ratios:

effects observed, treatment-related

Histopathological findings: non-neoplastic:

effects observed, non-treatment-related

Description (incidence and severity):

All findings occurred either individually or were biologically equally distributed over control and treatment groups. They were considered to be incidental or spontaneous in origin and without any relation to treatment.

Histopathological findings: neoplastic:

no effects observed

Reproductive function: oestrous cycle:

no effects observed

Description (incidence and severity):

Estrous cycle data, generated during the last 2 weeks prior to mating for the F1 litter, revealed regular cycles in the females of all test groups. The mean estrous cycle duration was similar: 3.9 / 3.9 / 3.9 and 4.0 days in test groups 0-3, respectively.

Reproductive function: sperm measures:

no effects observed

Description (incidence and severity):

The stages of spermatogenesis in the testes of males of the high dose test group were comparable to those of the controls. In high dose females the different stages of functional bodies in the ovaries were present and comparable to the control animals.

Reproductive performance:

effects observed, non-treatment-related

Description (incidence and severity):

Males
For all F0 parental males, which were placed with females to generate F1 pups, copulation was confirmed. Thus, the male mating index was 100% in all test groups.
Fertility was proven for all F0 parental males within the scheduled mating interval for F1 litter.
Thus, the male fertility index was 100% in all test groups.

Females
The female mating index calculated after the mating period for F1 litter was 100% in all test groups.
The mean duration until sperm was detected (GD 0) varied between 1.6 and 3.2 days without any relation to dosing. However, the low-dose value (3.2* days [p ≤ 0.05]) was statistically significantly increased in comparison to the concurrent control. This was mainly caused by the unusually low control value (1.6 days) which was even slightly below the historical control range (HCD: mean 2.6 [1.8-6.2] days) whereas the low-dose value was clearly within. Thus, and due to the lack of dose-response relationship, it is considered to be spontaneous in nature and not treatment related.
All female rats delivered pups or had implants in utero: the fertility index was 100% in all test groups.
The mean duration of gestation values varied between 22.2 / 22.7 / 22.4 / 22.3 in test groups 0-3, respectively.
The gestation index was 100% in test groups 0, 1 and 3. One mid-dose dam (No. 130) had only stillborn pups at delivery, thus, the gestation index was 90% in test group 2. Furthermore, the number of dams with stillborn pups was statistically significantly increased in test group 2 (4* [p ≤ 0.05] vs. 0 in control). This is considered as an incidental event, as there is no dose response relationship visible.
Implantation was not affected by the treatment since the mean number of implantation sites was comparable between all test substance-treated groups and the control, taking normal biological variation into account (13.1 / 13.2 / 13.6 and 13.5 implants/dam in test groups 0-3, respectively). Furthermore, there were no indications for test substance-induced intrauterine embryo-/fetolethality since the postimplantation loss did not show any significant differences between the groups, and the mean number of F1 pups delivered per dam remained unaffected (12.7 / 11.9 / 12.3 and 13.1 pups/dam in test groups 0-3, respectively).
The rate of liveborn pups was also not affected by the test substance, as indicated by live birth indices of 100.0% / 94.1% / 90.2% and 100% in test groups 0-3, respectively. Moreover, the number of stillborn pups was not significantly different between the test groups.
Thus, the test substance C13-C15-Alcohol did not adversely affect reproduction and delivery of the F0 generation parental females.

Key result

Dose descriptor:

NOAEL

Effect level:

1 000 mg/kg bw/day (nominal)

Based on:

test mat.

Sex:

male/female

Remarks on result:

not determinable due to absence of adverse toxic effects

Key result

Critical effects observed:

no

Clinical signs:

effects observed, non-treatment-related

Description (incidence and severity):

There were no test substance-related adverse clinical signs observed in any of the F1 generation pups of the different test groups.
Spontaneous findings occurred in test group 1:
• Male pup No. 114-02 had an injury and a swelling (due to this injury) on its left hindlimb on PND 12-13.
• Female pup No. 116-07 had a thread-like tail on PND 0-4.
• Four (out of 11) pups of dam No. 118 (with clinical observations, see 4.1.2.1.) were in a severe reduced nutritional condition (no milk in stomach).
Due to the lack of dose response relationship, these isolated findings are considered to be spontaneous in nature.

Dermal irritation (if dermal study):

not examined

Mortality / viability:

mortality observed, non-treatment-related

Description (incidence and severity):

Pup number and status at delivery
The mean number of delivered F1 pups per dam and the rates of liveborn and stillborn F1 pups were evenly distributed among the test groups. The respective values reflect the normal range of biological variation inherent in the strain used in this study.

Pup viability
The viability index indicating pup survival during lactation (PND 0-4) varied between 96.1% / 89.2% / 99.2% and 100% in test groups 0-3, respectively.
The pups surviving index indicating pup survival during lactation (PND 4-13) was 100% in all test groups.
Thus, the test substance did not influence pup survival in any of the treated groups (100, 300 and 1000 mg/kg bw/d).

Pup mortality
Pup mortality was not influenced by the test substance.

Body weight and weight changes:

effects observed, non-treatment-related

Description (incidence and severity):

In the high-dose group, the mean body weights of the female pups and, as a consequence, of both sexes combined were statistically significantly reduced on PND 13. Furthermore, the mean body weight changes in this test group were statistically significantly reduced in comparison to the concurrent control on PND 7-13 and 1-13.
As the values (PND 13 females 30.9 g) are fully covered by the historical control range (females 31.4 g [27.7-33.3 g] and the control values (day 13 females 33.5 g) are in this case outside the historical control range this perceived finding was assessed as incidental and not toxicological relevant.
The mean body weights and body weight change of all male and female pups in test groups 1 and 2 were comparable to the concurrent control values throughout the entire study.
One male runt, each, was seen in the control, the mid- and high-dose group. There were no female runts in any of the groups.

Food consumption and compound intake (if feeding study):

not examined

Food efficiency:

not examined

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

not examined

Ophthalmological findings:

not examined

Haematological findings:

not examined

Clinical biochemistry findings:

no effects observed

Description (incidence and severity):

Thyroid hormones
In parental males as well as in male and female pups at PND13 (test groups 1, 2 and 3; 100, 300 and 1000 mg/kg bw/d) no treatment-related alterations of T4 and TSH levels were observed.

Urinalysis findings:

not examined

Sexual maturation:

not examined

Anogenital distance (AGD):

no effects observed

Description (incidence and severity):

The anogenital distance and anogenital index of all test substance treated male and female pups was comparable to the concurrent control values.

Nipple retention in male pups:

no effects observed

Description (incidence and severity):

The apparent number and percentage of male pups having areolae was not influenced by the test substance when examined on PND 13.

Organ weight findings including organ / body weight ratios:

not examined

Gross pathological findings:

effects observed, non-treatment-related

Description (incidence and severity):

A few pups showed spontaneous findings at gross necropsy, such as:
• dilated renal pelvis in one male and four female control and two female mid-dose pups,
• discolored testis in two male mid-dose pups,
• a dilated ductus arteriosus in one female mid-dose pup,
• an empty stomach in three female low-dose pups,
• a thread-like tail in one female low-dose pup which was further assessed by skeletal examination (sacral vertebrae misshapen or absent, all caudal vertebrae absent).
These findings occurred without any relation to dosing. Thus, all these findings were considered to be not associated to the test substance.

Histopathological findings:

not examined

Other effects:

effects observed, non-treatment-related

Description (incidence and severity):

Sex ratio
The sex distribution and sex ratios of live F1 pups on the day of birth and PND 13 did not show substantial differences between the control and the test substance-treated groups; slight differences were regarded to be spontaneous in nature.

Behaviour (functional findings):

not examined

Developmental immunotoxicity:

not examined

Key result

Dose descriptor:

NOAEL

Generation:

F1

Effect level:

1 000 mg/kg bw/day (nominal)

Based on:

test mat.

Sex:

male/female

Remarks on result:

not determinable due to absence of adverse toxic effects

Key result

Critical effects observed:

no

Key result

Reproductive effects observed:

no

Conclusions:

In conclusion, under the conditions of the present OECD 422 combined repeated dose toxicity study with the reproductive/developmental screening test in Wistar rats the oral administration of C13-C15-Alcohol by gavage to male and female Wistar rats resulted in no signs of systemic toxicity up to a dose of 1000 mg/kg bw/d.
Thus, the no observed adverse effect level (NOAEL) for general systemic toxicity was 1000 mg/kg bw/d for male and female Wistar rats.
The NOAEL for reproductive performance and fertility was set to 1000 mg/kg bw/d for male and female Wistar rats.
The NOAEL for developmental toxicity was 1000 mg/kg bw/d.

Executive summary:

In an OECD 422 study, the test substance C13-C15-Alcohol was administered daily to groups of 10 male and 10 female Wistar rats (F0 animals) by gavage at doses of 100, 300 and 1000 mg/kg body weight/day (mg/kg bw/d, test groups 1-3, respectively) to screen for potential systemic, reproductive and developmental toxicity. Control animals (10 male and 10 female Wistar rats) were dosed daily with the vehicle only (0.5% Sodium carboxymethyl cellulose suspension (CMC) in deionized water with 10 mg/ 100 mL Tween 80). The duration of treatment covered a 37 days in-life period in males (including premating, mating [mating pairs were from the same test group] and postmating period) and a 2-weeks premating and mating period, the entire gestation and approximately 3 weeks of lactation period in females. Parental females were allowed to give birth and bring up the offspring until sacrifice on PND 4 or 13.
The stability of these preparations was demonstrated over a period of 7 days under ambient conditions. Analyses confirmed the overall accuracy of the prepared concentrations and the homogeneity of the test substance in the vehicle.
Regarding clinical examination, no test substance related, adverse signs of (systemic) toxicity were observed up to limit dose of 1000 mg/ kg bw/d.
Most of the high- and mid-dose and single low-dose animals showed temporary salivation after dosing during the entire study. Salivation occurred immediately after dosing (up to 2 hours post dosing), and also afterwards (between 2 and 5 hours post dosing). It is most likely, that this temporary finding was induced by a bad taste of the test substance or local affection of the upper digestive tract. It is, however, not considered to be an adverse toxicologically relevant finding.
Neither body weight development nor food consumption were adversely affected in any of the tested dose groups.
Concerning clinical pathology, no treatment-related, adverse effects were observed up to a dose of the compound of 1000 mg/kg bw/d.
Regarding pathology, target organ was the liver in female animals of test group 3 which showed an increased mean relative weight. This was considered possibly treatment-related, but not adverse due to the absence of histopathological findings.
Regarding fertility and reproductive performance, no signs of toxicity were observed in male or female parental animals of all test groups during the entire study. F0 parental animals proved to be fertile. Mating behavior, conception, implantation and parturition were not affected.
Regarding developmental toxicity, no signs of developmental toxicity were noted in any of the treated groups. Pup status, viability, survival and growth showed no treatment-related, adverse findings.
All other findings occurred either individually or were biologically equally distributed over control and treatment groups. They were considered to be incidental or spontaneous in origin and without any relation to treatment.

Effect on fertility: via oral route

Endpoint conclusion:

no adverse effect observed

Dose descriptor:

NOAEL

1 000 mg/kg bw/day

Species:

rat

Additional information

In an OECD 422 study, the test substance C13-C15-Alcohol was administered daily to groups of 10 male and 10 female Wistar rats (F0 animals) by gavage at doses of 100, 300 and 1000 mg/kg body weight/day (mg/kg bw/d, test groups 1-3, respectively) to screen for potential systemic, reproductive and developmental toxicity. Control animals (10 male and 10 female Wistar rats) were dosed daily with the vehicle only (0.5% Sodium carboxymethyl cellulose suspension (CMC) in deionized water with 10 mg/ 100 mL Tween 80). The duration of treatment covered a 37 days in-life period in males (including premating, mating [mating pairs were from the same test group] and postmating period) and a 2-weeks premating and mating period, the entire gestation and approximately 3 weeks of lactation period in females. Parental females were allowed to give birth and bring up the offspring until sacrifice on PND 4 or 13. The stability of these preparations was demonstrated over a period of 7 days under ambient conditions. Analyses confirmed the overall accuracy of the prepared concentrations and the homogeneity of the test substance in the vehicle. Regarding clinical examination, no test substance related, adverse signs of (systemic) toxicity were observed up to limit dose of 1000 mg/ kg bw/d. Most of the high- and mid-dose and single low-dose animals showed temporary salivation after dosing during the entire study. Salivation occurred immediately after dosing (up to 2 hours post dosing), and also afterwards (between 2 and 5 hours post dosing). It is most likely, that this temporary finding was induced by a bad taste of the test substance or local affection of the upper digestive tract. It is, however, not considered to be an adverse toxicologically relevant finding. Neither body weight development nor food consumption were adversely affected in any of the tested dose groups. Concerning clinical pathology, no treatment-related, adverse effects were observed up to a dose of the compound of 1000 mg/kg bw/d. Regarding pathology, target organ was the liver in female animals of test group 3 which showed an increased mean relative weight. This was considered possibly treatment-related, but not adverse due to the absence of histopathological findings. Regarding fertility and reproductive performance, no signs of toxicity were observed in male or female parental animals of all test groups during the entire study. F0 parental animals proved to be fertile. Mating behavior, conception, implantation and parturition were not affected. Regarding developmental toxicity, no signs of developmental toxicity were noted in any of the treated groups. Pup status, viability, survival and growth showed no treatment-related, adverse findings. All other findings occurred either individually or were biologically equally distributed over control and treatment groups. They were considered to be incidental or spontaneous in origin and without any relation to treatment.

Effects on developmental toxicity

Description of key information

No adverse effects on embryotoxicity or teratogenicity was observed with the two structural analoques.

Effect on developmental toxicity: via oral route

Endpoint conclusion:

no adverse effect observed

Dose descriptor:

NOAEL

750 mg/kg bw/day

Species:

rat

Additional information

The potential of alcohols C13 - 15 branched and linear to induce developmental toxicity was deduced from the structural analogues with CAS numbers 27458 -92 -0 (BASF SE, 2003 & 2020) 10042 -59 -8 (BASF SE, 2004).

Developmental toxicity of CAS 27458-92-0 was examined in a GLP-study (according to OECD 414), where 25 Wistar rats (per dose) were administered 60, 250 and 750 mg/kg bw of test material orally via gavage. The administration was daily from day 6 through day 19 post coitum. On gestation day 20, the dams were sacrificed and ovaries and uterine content as well as fetuses were examined.There were no substance-related or spontaneous mortalities in any of the groups. Some signs of maternal toxicity at 750 mg/kg body weight/day were detected. Clinically, transient salivation (in all rats), urine smeared fur (in 4 dams), and reduced food consumption at initiation of treatment was observed at the high dose level. Clinical pathology revealed increased alanine aminotransferase activities, decreased total protein and globulin concentrations, and increased triglyceride levels in the serum of the high dose females. These changes, which were accompanied by statistically significantly increased absolute and relative liver weights (about 14% or 18% respectively above control values), are indicative for a mild adverse effect on the liver. The only substance-induced finding on the mid dose dams (250 mg/kg body weight/day) consisted in transitory salivation in 17 out of 25 rats, which, by itself and if seen in isolation, is not assessed as an adverse or toxic effect. No substance-induced effects on the dams occurred at the low dose level (60 mg/kg body weight/day). The oral administration of the test substance to the dams at all 3 dose levels (60, 250 and 750 mg/kg body weight/day) had no influence on the gestational parameters and evoked no signs of prenatal developmental toxicity and in particular no indications for teratogenicity at the 3 dose levels tested. Placental and fetal body weights were unaffected and the external, soft tissue and/or skeletal (including cartilage) examinations of the fetuses revealed no biologically relevant differences between the control and the substance-treated groups (i.e. 60, 250 and 750 mg/kg body weight/day). Based on these results, the NOAEL for maternal toxicity is 250 mg/kg body weight/day, while it is 750 mg/kg body weight/day for prenatal developmental toxicity.

Additionally, the developmental toxicity of test substance CAS 10042-59-8 wasexamined in a reliable GLP-study (according to OECD 414), where 25 Wistar rats (per dose) were administered an aqueous emulsion of 50, 200 and 600 mg/kg bw test material orally via gavage. The administration was daily from day 6 through day 19 post coitum. On gestation day 20, the dams were sacrificed and ovaries and uterine content as well as fetuses were examined. At 600 mg/kg bw signs of maternal toxicity like clinically salivation and discomfort, reduced food consumption, reduced absolute and net body weight gain and increased water consumption were recorded. Clinical pathology revealed slight changes such as mild thrombocytopenia and marginal changes in electrolyte levels in the peripheral blood of the high dose dams. Organ weight determination revealed an increased absolute and relative liver weight. Also, at mid dose level (200 mg/kg bw) were still some signs of maternal toxicity such as salivation, reduced absolute and net maternal body weight gain as well as a slightly increased relative liver weight recorded (NOAEL for maternal toxicity = 50 mg/kg bw). The increase in the absolute and relative liver weight (200 and 600 mg/kg bw) is likely to be associated to a peroxisome proliferation and has therefore no human hazard. The oral administration of the test material to dams at all 3 dose levels had no influence on gestational parameters. Conception rate, mean number of corpora lutea, total implantations, resorptions and live fetuses, fetal sex ratio or values calculated for pre- and postimplantation losses were unaffected by the treatment. There was a statistically significant reduction of mean fetal body weight at the high dose level (600 mg/kg bw) of about 11% below the corresponding control value. Since this value was still in the range of historical control data, the finding is assessed to be secondary to the distinct maternal toxicity that has been observed in the appropriate dams. The mean fetal body weights in low and mid dose (50 and 200 mg/kg bw) were not influenced by the test substance. No toxicologically relevant influence of the test substance on mean placental weights was noted. The external, soft tissue and/or skeletal examinations of the fetuses revealed no malformations which might be related to the test substance and no specific malformation pattern was found. A slight increase of the overall incidence of skeletal variations was noted at high dose level (600 mg/kg bw), but also these findings are considered as secondary to maternal toxicity and not relevant in terms of developmental toxicity. Therefore the NOAEL for embryotoxicity/teratogenicity and fetotoxicity was 600 mg/kg bw.

Further, CAS 27458-92-0 was administered to 25 pregnant New Zealand White rabbits (per dose) in a prenatal developmental toxicity study (according to OECD 414), daily by stomach tube from implantation to one day prior to the expected day of parturition (GD 6-28). Clinical observations and records of food consumption and body weight/body weight gain revealed no toxicologically relevant difference between the animals receiving 25 or 75 mg/kg bw/d Isotridecanol and the controls. The high dose of 250 mg/kg bw/d produced a decrease of food consumption as well as body weight/body weight gain during the first two of three treatment weeks. Although the animals partly compensated the lower food intake in the last treatment week, the high-dose does consumed overall 8% less food and had peak food consumption decreases of 31% compared to the concurrent control does during the treatment period (GD 6-28). Accordingly, lower food intake resulted in weight loss (-16.6 g vs. 28.4 g in the control) at the beginning of treatment, and the high-dose body weight remained below control during most of the treatment period. Only towards the end of treatment the high-dose mean body weights recovered and were comparable to the control again. If calculated for the treatment period (GD 6-28), the high-dose does gained about 8% less weight in comparison to the control does (without attaining statistical significance). Concerning clinical pathology, no treatment-related, adverse effects were observed up to a dose of the test compound of 250 mg/kg bw/d. Concerning pathology, significantly increased absolute and relative kidney weights in test groups 2 and 3 as well as absolute and relative liver weights in test group 3 were regarded as treatment-related. 

There were no test substance-related and/or biologically relevant differences between thedifferent test groups in conception rate, in the mean numbers of corpora lutea and implantation sites or in the values calculated for the pre- and the post-implantation losses, the numbers of resorptions and viable fetuses. Similarly, no influence of the test substance on uterine weight, placental weight, fetal weight and sex distribution of the fetuses was noted at any dose. All differences observed are considered to reflect the normal range of fluctuations for animals of this strain and age. Fetal examinations revealed no toxicologically relevant adverse effects of the test substance on embryofetal development.

Under the conditions of this prenatal developmental toxicity study, the oral administration of Isotridecanol to pregnant New Zealand White rabbits from implantation to one day prior to the expected day of parturition (GD 6-28) caused evidence of systemic maternal toxicity at the high-dose level of 250 mg/kg bw/d, such as decrease of food consumption and body weight/body weight gain during major parts of the treatment. In conclusion, the no observed adverse effect level (NOAEL) for maternal toxicity is 75 mg/kg bw/d.As there was no evidence of toxicologically relevant adverse effects of the test substance on embryofetal development, the no observed adverse effect level (NOAEL) for prenatal developmental toxicity is 250 mg/kg bw/d.

Justification for classification or non-classification

Based on the available information the substance does not need to be classified for developmental toxicity, as in accordance with EU Classification, Labeling and Packaging of Substances and Mixtures (CLP) Regulation No. 1272/2008.