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Description of key information

Oral: OECD 422, rat, NOAEL ≥ 1000 mg/kg bw/day

Key value for chemical safety assessment

Repeated dose toxicity: via oral route - systemic effects

Link to relevant study records
Reference
Endpoint:
short-term repeated dose toxicity: oral
Remarks:
combined repeated dose and reproduction / developmental screening
Type of information:
migrated information: read-across from supporting substance (structural analogue or surrogate)
Adequacy of study:
key study
Study period:
06 Jun - 04 Aug 2013
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: see 'Remark'
Remarks:
GLP - guideline study, tested with the source substance tetradecyl oleate (CAS 22393-85-7). According to the ECHA guidance document 'Practical guide 6: How to report read-across and categories (ECHA, 2012)', the reliability was changed from RL1 to RL2 to reflect the fact that this study was conducted on a read-across substance.
Qualifier:
according to guideline
Guideline:
OECD Guideline 422 (Combined Repeated Dose Toxicity Study with the Reproduction / Developmental Toxicity Screening Test)
Version / remarks:
adopted in 22 Mar 1996
Deviations:
no
GLP compliance:
yes
Limit test:
no
Species:
rat
Strain:
other: Crl:CD(SD)BR
Sex:
male/female
Details on test animals or test system and environmental conditions:
TEST ANIMALS
- Source: Charles River Italia S.p.A., Calco (Lecco), Italy
- Age at study initiation: 6 to 7 weeks
- Weight at study initiation: 191 to 204 g (males) and 172 to 179 g (females)
- Housing: From arrival to pairing: animals were housed 5 of one sex to a cage, in polysulphone solid bottomed cages measuring 59.5x38x20 cm (Techniplast Gazzada S.a.r.l., Buguggiate, Varese, Italy). Nesting material was provided inside suitable bedding bags and changed at least twice a week.
During mating: animals were housed one male to one female in clear polysulphone cages measuring approximately 43x27x18 cm with a stainless steel mesh lid and floor (Techniplast Gazzada S.a.r.l., Buguggiate, Varese, Italy). Each cage tray held absorbent material which was inspected and changed daily. After mating, the males were re-caged as they were before mating; the females were transferred to individual solid bottomed cages (Techniplast Gazzada S.a.r.l., Buguggiate, Varese, Italy) for the gestation period and parturition.
- Diet: laboratory rodent diet, 4 RF 21 (Mucedola S.r.l., Settimo Milanese (MI), Italy), ad libitum
- Water: drinking water, ad libitum
- Acclimation period: 2 weeks

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22 ± 2
- Humidity (%): 55 ± 15
- Air changes (per hr): 15-20
- Photoperiod (hrs dark / hrs light): 12/12
Route of administration:
oral: gavage
Vehicle:
other: 0.5% aqueous carboxymethylcellulose (0.5% CMC)
Details on oral exposure:
PREPARATION OF DOSING SOLUTIONS: The test substance was suspended in the vehicle. Formulations were prepared daily.

VEHICLE
- Concentration in vehicle: 10, 30 and 100 mg/mL for dose levels of 100, 300 and 1000 mg/kg bw/day, respectively
- Amount of vehicle: 10 mL/kg bw
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
Concentration, homogeneity and stability of the test substance in the vehicle were verified by gas chromatopgraphy with a flame ionisation detection (FID). Concentration verification was conducted on a weekly basis.
Duration of treatment / exposure:
Males: The daily administration of the test item was started two weeks before mating and lasted until test day 28 to 29, which was one day before sacrifice.
Females: The daily administration of the test item was started two weeks before mating and continued until day 3 post-partum.
Maximum: 54 days of treatment.
Frequency of treatment:
once daily, 7 days/week
Remarks:
Doses / Concentrations:
100, 300 and 1000 mg/kg bw/day
Basis:
actual ingested
No. of animals per sex per dose:
10
Control animals:
yes, concurrent vehicle
Details on study design:
- Dose selection rationale: based on a 14-day range-finding study (Rossiello, 2013. RTC Study No.: 93730EXT)
Observations and examinations performed and frequency:
CAGE SIDE OBSERVATIONS: Yes
- Time schedule: at least once daily during the study, each animal was observed and any clinical signs recorded.

DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: once before commencement of treatment and at least once a week thereafter, each animal was given a detailed clinical examination. Each animal was removed from the home cage and observed in an open arena. The tests included observation of changes in gait and posture, reactivity to handling, presence of clonic or tonic movements, stereotypes or bizarre behaviour and effects on the autonomic nervous system (e.g. lachrymation, piloerection, pupil size, unusual respiratory pattern).

BODY WEIGHT: Yes
- Time schedule for examinations: females: weekly from allocation to positive identification of mating and on gestation days 0, 7, 14 and 20. Dams were also weighed on Day 1 and 4 post partum.

FOOD CONSUMPTION AND COMPOUND INTAKE
- Food consumption for each animal determined and mean daily diet consumption calculated as g food/kg body weight/day: Yes. The weight of food consumed by each cage of males and females was recorded weekly during the pre-mating period starting from allocation. Individual food consumption for the females was measured on gestation days 7, 14 and 20 starting from Day 0 post coitum and on Day 4 post partum starting from Day 1 post partum.
- Compound intake calculated as time-weighted averages from the consumption and body weight gain data: No data

FOOD EFFICIENCY:
- Body weight gain in kg/food consumption in kg per unit time X 100 calculated as time-weighted averages from the consumption and body weight gain data: No

WATER CONSUMPTION AND COMPOUND INTAKE : Yes
- Time schedule for examinations: daily by visual appraisal

OPHTHALMOSCOPIC EXAMINATION: No

HAEMATOLOGY: Yes
- Time schedule for collection of blood: at the end of treatment
- Anaesthetic used for blood collection: Yes with isofluorane
- Animals fasted: Yes
- How many animals: 5/sex/dose
- Parameters examined: haematocrit, haemoglobin, red blood cell count, reticulocyte count, mean red blood cell volume, mean corpuscular haemoglobin, mean corpuscular haemoglobin concentration, white blood cell count and differential leucocyte count

Coagulation tests: Prothrombin time

CLINICAL CHEMISTRY: Yes
- Time schedule for collection of blood: at the end of treatment
- Animals fasted: Yes
- How many animals :5/sex/dose
- Parameters examined: albumin, globulin, albumin/globulin ratio, bile acids, total bilirubin, total cholesterol, creatinine, glucose, urea, total protein, calcium, chloride, potassium, sodium, alanine aminotransferase (ALAT), alkaline phosphatase (AP), aspartate aminotransferase (ASAT), gamma-glutamyltransferase, triglycerides, inorganic phosphorus

URINALYSIS: No

NEUROBEHAVIOURAL EXAMINATION: Yes
- Time schedule for examinations: once during the study, towards the end of treatment, 5 males and 5 females were randomly
selected from each group for evaluation of sensory reactivity to stimuli of different modalities (e.g. auditory, visual and proprioceptive stimuli), for assessment of grip strength and for the motor activity was measured (for approximately 5 min).
- Dose groups that were examined: all dose groups
- Battery of functions tested: sensory reactivity to stimuli of different types (e.g. auditory, visual and proprioceptive stimuli) (based on Gad), as well as the assessment of grip strength (Meyer) and motor activity

Sacrifice and pathology:
GROSS PATHOLOGY: Yes
-Organ weights: adrenal glands, brain (cerebrum, cerebellum, medulla/pons), epididymides, heart, kidneys, liver, ovaries with oviducts, parathyroid glands, prostate gland, seminal vesicles with coagulating glands, spleen, testes, thymus (where present), thyroid and uterus-cervix,
-Fixation: adrenal glands, bone marrow (from sternum), brain (cerebrum, cerebellum, medulla/pons), caecum, clitoral gland, colon, duodenum, epididymides, heart, ileum (including Peyer’s patches), jejunum, kidneys, liver, lungs (including mainstem bronchi), lymph nodes (mesenteric and cervical), ovaries with oviducts, parathyroid glands, pituitary gland, penis, preputial gland, prostate gland, rectum, sciatic nerve, seminal vesicles with coagulating glands, spinal column, spinal cord (cervical, thoracic, lumber), spleen, stomach, testes, thymus (where present), thyroid, trachea, urinary bladder, uterus-cervix and vagina.

HISTOPATHOLOGY: Yes, all organs that were included for fixation (5/sex of control and high dose group)

In addition, the testes and epididymides were cut at 2-3 micrometer thickness and stained with Periodic Acid Schiff (PAS). The morphological evaluation of the seminiferous epithelium (staging of spermatogenic cycle) was performed. A detailed qualitative evaluation of testes was performed on 5 randomly selected control and high dose males. The evaluation took into account the tubular stages of the spermatogenic cycle, in order to identify treatment-related effects such as: missing germ cell layers or types, retained spermatids, multinucleated or apoptotic germ cells and sloughing of spermatogenic cells into the lumen.
Other examinations:
Vaginal smears were taken daily in the morning starting two weeks before pairing until a positive identification of copulation was made.
Statistics:
Standard deviations were calculated as appropriate. For continuous variables the significance of the differences amongst group means was assessed by Dunnett’s test or a modified t test, depending on the homogeneity of data. The non-parametric Kruskal-Wallis analysis of variance was used for the other parameters. Intergroup differences between the control and treated groups were assessed by the nonparametric version of the Williams test.
The criterion for statistical significance was p<0.05.
Clinical signs:
no effects observed
Mortality:
no mortality observed
Body weight and weight changes:
no effects observed
Food consumption and compound intake (if feeding study):
no effects observed
Food efficiency:
not examined
Water consumption and compound intake (if drinking water study):
no effects observed
Ophthalmological findings:
not examined
Haematological findings:
no effects observed
Clinical biochemistry findings:
no effects observed
Urinalysis findings:
not examined
Behaviour (functional findings):
no effects observed
Organ weight findings including organ / body weight ratios:
no effects observed
Gross pathological findings:
no effects observed
Histopathological findings: non-neoplastic:
no effects observed
Histopathological findings: neoplastic:
not examined
Details on results:
CLINICAL SIGNS AND MORTALITY
No relevant clinical signs or mortality were observed in males and females throughout the study.

BODY WEIGHT AND WEIGHT GAIN
No differences of toxicological significance were seen in terminal body weight or body weight gain.

FOOD CONSUMPTION AND COMPOUND INTAKE
No intergroup differences were seen in food consumption.

HAEMATOLOGY
No changes of toxicological significance were observed. The statistically significant differences between control and treated animals (erythrocytes, mean corpuscular haemoglobin and leucocytes in males, haemoglobin, haematocrit and platelets in females) were of low magnitude and/or not dose-related, therefore considered incidental.

CLINICAL CHEMISTRY
No changes of toxicological significance were observed. Statistically significant fluctuations of some biochemical parameters (mean group values) were recorded such as: increase of glucose in males dosed with 100 and 1000 mg/kg bw/day (48% for both dosages), increase in urea in those receiving 100 mg/kg bw/day (19%), increase of aspartate aminotransferases in females dosed with 300 mg/kg bw/day (35%), decrease of bilirubin (81%) and increase of potassium (10%) in those treated with 1000 mg/kg/day when compared to controls.
Due to the lack of dose- and sex-consistency and to the absence of other relevant findings, the above alterations were considered unrelated to treatment.


NEUROBEHAVIOUR
No significant differences were observed between groups, in motor activity, grip strength and sensory reactivity to stimuli, evaluated at the end of treatment.

ORGAN WEIGHTS
No relevant differences in organ weights were seen between the controls and treated animals of both sexes.

GROSS PATHOLOGY
No remarkable changes were noted at post mortem examination in treated animals when compared with controls.

HISTOPATHOLOGY
No treatment-related changes were observed. The lesions reported in control and treated animals were considered to be an expression of spontaneous and/or incidental pathology, commonly seen in this species and age under the experimental conditions.

OTHER FINDINGS:
Spermatogenic cycle
Seminiferous tubules were evaluated with respect to their stage in the spermatogenic cycle and to the integrity of the various cell types within the different stages; regular layering in the germinal epithelium was noted. No relevant difference in estrous cycle was observed in treated females when compared to controls.
Dose descriptor:
NOAEL
Effect level:
>= 1 000 mg/kg bw/day (actual dose received)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
other: no adverse effects observed
Critical effects observed:
not specified
Endpoint conclusion
Endpoint conclusion:
no adverse effect observed
Quality of whole database:
The available information comprises adequate, reliable (Klimisch score 2 due to read-across) studies from reference substances with similar structure and intrinsic properties. Read-across is justified based on common precursors and breakdown products of hydrolysis and consistent trends in environmental fate, ecotoxicological and toxicological profile (refer to the endpoint discussion for further details).
The selected study is thus sufficient to fulfil the standard information requirements set out in Annex VIII, 8.6, in accordance with Annex XI, 1.5, of Regulation (EC) No 1907/2006.

Repeated dose toxicity: inhalation - systemic effects

Endpoint conclusion
Endpoint conclusion:
no study available

Repeated dose toxicity: inhalation - local effects

Endpoint conclusion
Endpoint conclusion:
no study available

Repeated dose toxicity: dermal - systemic effects

Endpoint conclusion
Endpoint conclusion:
no study available

Repeated dose toxicity: dermal - local effects

Endpoint conclusion
Endpoint conclusion:
no study available

Additional information

Justification for read-across

There are no available data on the repeated dose toxicity of 2-octyldodecyl myristate (CAS 22766-83-2). The assessment was therefore based on studies conducted with analogue substances as part of a read across approach, which is in accordance with Regulation (EC) No. 1907/2006, Annex XI, 1.5. For each specific endpoint the source substance(s) structurally closest to the target substance is/are chosen for read-across, with due regard to the requirements of adequacy and reliability of the available data. Structural similarities and similarities in properties and/or activities of the source and target substance are the basis of read-across. A detailed justification for the analogue read-across approach is provided in the technical dossier (see IUCLID Section 13) and within Chapter 5.1 of the CSR.

Repeated dose toxicity, oral, subacute

CAS 22393-85-7

A 14-day dose range-finding study was performed with tetradecyl oleate (CAS 22393-85-7) in order to find appropriate dose levels for a subsequent combined repeated dose toxicity and reproduction/developmental toxicity screening study according to OECD Guideline 422 (Rossiello, 2013). In this study, 5 rats per sex and dose were administered the test substance once daily at 100, 300 and 1000 mg/kg bw/day via oral gavage for 14 days. No mortality occurred and no clinical signs were observed during the study period. The body weight and food consumption was comparable between the control and treatment groups. Minor changes were observed in organ weights as noted by decreases in the absolute and relative thymus weight (approximately -18%) in males receiving 1000 mg/kg bw/day and statistically significant decreases in absolute and relative kidneys weight (-12%, for absolute weights) in females receiving 1000 mg/kg bw/day. These effects were not considered to be adverse, as they were observed in one sex only and no corresponding histopathological changes were seen. Based on the results of this study, the subacute NOAEL for male and female rats was considered to be ≥ 1000 mg/kg bw/day.

A combined repeated dose toxicity and reproduction/developmental toxicity screening study (according to OECD Guideline 422 and in compliance with GLP) was performed (Rossiello, 2014). 10 rats/sex/dose were administered 0, 100, 300 and 1000 mg/kg bw/day tetradecyl oleate once daily for 28-29 days (males) and up to 54 days (females) via gavage. The application started two weeks before mating on test day one and ended on the day of or one day before sacrifice. Day of sacrifice was on test day 29 or 30 for the male rats and on lactation day 3 or shortly thereafter for the female rats. There was no mortality during the study period. No toxicologically relevant clinical signs were observed. The body weight, body weight gain and food consumption was comparable between the control and treatment groups. There were no toxicologically relevant effects on organ weights. The statistically significant differences in haematological parameters between control and treated animals (erythrocytes, mean corpuscular haemoglobin and leucocytes in males, haemoglobin, haematocrit and platelets in females) were of low magnitude and/or not dose-related, and therefore considered incidental. Statistically significant fluctuations of some biochemical parameters, compared with the control groups, were recorded in both sexes. An increase in glucose levels in males administered 100 and 1000 mg/kg bw/day (48% for both doses), and an increase in urea in males receiving 100 mg/kg bw/day (19%) was observed. In females in the 300 mg/kg bw/day group, an increase in aspartate aminotransferase level (35%) was noted, while for females administered 1000 mg/kg bw/day, a decrease in bilirubin levels (81%) and an increase in potassium levels (10%) were observed. Due to the lack of dose- and/or sex-consistency, and due to the absence of other relevant findings, these changes are not considered to be toxicologically relevant. There was no significant difference between control and treatment groups during the observational and neurological screenings. The macroscopic inspection at autopsy and subsequent histopathological examination did not reveal any treatment-related changes. The NOAEL for systemic toxicity was considered to be ≥ 1000 mg/kg bw/day.

 

CAS 93803-87-3

A 28-day oral repeated dose toxicity study was performed according to OECD 407, using octyldodecyl isooctadecanoate (De Hoog, 1998). 5 Wistar rats/sex/dose were administered 50, 200 and 1000 mg/kg bw/day by gavage, for 28 consecutive days. There was no mortality and no toxicologically relevant clinical signs were observed during the study period. No significant differences in body weight, body weight gain and food consumption between the control group and treatment groups were noted. In females of the high-dose group, a statistically significant increase in relative leucocyte level (approx. 8%) and decrease in relative neutrophil level (approx. 8%) was noted, compared with the control group values. These values are related to each other as a percentage of the total lymphocyte concentration, and a variation in one will necessarily affect one or several other specific lymphocyte values. As the changes are minimal, no effects were seen in males and no other haematological effects were noted, this result is considered not have toxicological relevance. In males of the high-dose group, the level of calcium and chloride was significantly increased. These increases were around 5% compared with the control group levels and no similar effects were observed in the female groups. Therefore, they are not considered to have toxicological relevance. No treatment-related effects on neurobehavioural parameters were observed. There were no significant differences in absolute or relative organ weight in the treatment groups, compared to the control group. The macroscopic inspection at autopsy and subsequent histopathological examination did not reveal any treatment-related changes. Based on the lack of toxicologically relevant effects up to and including the highest dose level, the NOAEL is considered to be ≥ 1000 mg/kg bw/day.

 

 

Overall conclusion for repeated dose toxicity

The data for the read-across analogue substances showed that no effects were observed up to and including the recommended limit value. Therefore, as the available data did not identify any hazard for repeated dose toxicity, 2-octyldodecyl myristate is not considered to be hazardous following repeated exposure.


Justification for selection of repeated dose toxicity via oral route - systemic effects endpoint:
Hazard assessment is conducted by means of read-across from a structural analogue. The selected study is the most adequate and reliable study based on the identified similarities in structure and intrinsic properties between the source and target substance and overall assessment of quality, duration and dose descriptor level (refer to the endpoint discussion for further details).

Justification for classification or non-classification

According to Article 13 of Regulation (EC) No. 1907/2006 "General Requirements for Generation of Information on Intrinsic Properties of substances", information on intrinsic properties of substances may be generated by means other than tests e.g. from information from structurally related substances (grouping or read-across), provided that conditions set out in Annex XI are met. Annex XI, "General rules for adaptation of this standard testing regime set out in Annexes VII to X” states that “substances whose physicochemical, toxicological and ecotoxicological properties are likely to be similar or follow a regular pattern as a result of structural similarity may be considered as a group, or ‘category’ of substances. This avoids the need to test every substance for every endpoint". Since the analogue concept is applied to 2-octyldodecyl myristate (CAS 22766-83-2), data will be generated from data for reference source substance(s) to avoid unnecessary animal testing. Additionally, once the analogue read-across concept is applied, substances will be classified and labelled on this basis.

Therefore, based on the analogue read-across approach, the available data on repeated dose toxicity do not meet the classification criteria according to Regulation (EC) 1272/2008 or Directive 67/548/EEC, and are therefore conclusive but not sufficient for classification.