Fatty acids, coco, esters with 1,3-butanediol

Repeated dose toxicity: oral

Administrative data

Endpoint:

sub-chronic toxicity: oral

Type of information:

experimental study

Adequacy of study:

key study

Study period:

16 Nov 2000 to 30 Nov 2001

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

comparable to guideline study

Remarks:

No guideline mentioned in study report

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2001

Report date:

2001

Materials and methods

Test guideline

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 408 (Repeated Dose 90-Day Oral Toxicity Study in Rodents)

Version / remarks:

adopted in 1998

Deviations:

yes

Remarks:

no immunological effects addressed

Principles of method if other than guideline:

No guideline mentioned in study report

GLP compliance:

yes

Limit test:

no

Test material

Test material information

Constituent 1

Reference substance name:

853947-59-8

Cas Number:

853947-59-8

IUPAC Name:

853947-59-8

Test animals

Species:

rat

Strain:

Sprague-Dawley

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Harlan ltaly S.r.l., 33049 San Pietro al Natisone (UD), Italy
- Females (if applicable) nulliparous and non-pregnant: not stated
- Age at study initiation: 27-29 days old
- Weight at study initiation: 89.0 - 102.1 g for males and 84.6 - 101.7 g for females
- Fasting period before study: none
- Housing: 5 of one sex to a cage, clear polycarbonate cages measuring 59x38.5x20 cm with a stainless steel mesh lid and floor (Code 1354 G, Techniplast Gazzada S.a.r.1., Buguggiate, Varese). Each cage tray held absorbent paper which was inspected daily and changed three times a week.
- Diet: Altromin MT pelleted diet, A. Rieper, Bolzano, ltaly, ad libitum
- Water: via water bottles, ad libitum
There was no information indicating that any non-nutrient substance likely to influence the effect of the test item was present in the drinking water or in the diet.
- Acclimation period: 13 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22 ± 2
- Humidity (%): 55 ± 15
- Air changes (per hr): 15-20
- Photoperiod (hrs dark / hrs light): 12/12

IN-LIFE DATES: 16 Nov 2000 to 22 Feb 2001

Administration / exposure

Route of administration:

oral: gavage

Details on route of administration:

Dose volume of 2 mL/kg body weight

Vehicle:

corn oil

Details on oral exposure:

PREPARATION OF DOSING SOLUTIONS: The required amount of the test item was dissolved in the vehicle (corn oil). The formulations were prepared daily. The dose was administered to each animal on the basis of the most recently recorded body weight, and the volume administered was recorded for each animal.

VEHICLE
- Justification for use and choice of vehicle (if other than water): not given
- Concentration in vehicle: 125, 250 and 500 mg/mL
- Lot/batch no. (if required): not given
- Purity: not given

Analytical verification of doses or concentrations:

no

Details on analytical verification of doses or concentrations:

None

Duration of treatment / exposure:

13 weeks

Frequency of treatment:

once daily

Doses / concentrations

Doses / concentrations 1

Dose / conc.:

0 mg/kg bw/day (nominal)

Doses / concentrations 2

Dose / conc.:

250 mg/kg bw/day (nominal)

Doses / concentrations 3

Dose / conc.:

500 mg/kg bw/day (nominal)

Doses / concentrations 4

Dose / conc.:

1 000 mg/kg bw/day (nominal)

No. of animals per sex per dose:

10

Control animals:

yes, concurrent vehicle

Details on study design:

- Dose selection rationale: not given

Positive control:

None

Examinations

Observations and examinations performed and frequency:

CAGE SIDE OBSERVATIONS: Yes
- Time schedule: For mortality twice a day

DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: Pre and post-dose observations of individual animals for signs of reaction to treatment was carried out daily before dosing, approximately 30 minutes and 2 hours after dosing. Once before commencement of treatment and once a week thereafter each animal was subjected to a detailed clinical examination, which included an evaluation of neurotoxicity. Animals were examined in an open arena for a period of three minutes: removal (from cage), handling reactivity, lacrymation, palpebral closure, salivation, piloerection, rearing, motility impairment, arousal (animal activity), vocalisation, stereotypies, unusual respiratory pattern, bizarre behaviour, urination, defecation, clonic and tonic movements, and gait.

BODY WEIGHT: Yes
- Time schedule for examinations: on the day of allocation to treatment groups, on the day that treatment commenced, weekly thereafter and just prior to necropsy.

FOOD CONSUMPTION:
The weight of food consumed by each cage of rats was recorded weekly following allocation and the group mean daily intake per rat was calculated.

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 (if drinking water study): No

OPHTHALMOSCOPIC EXAMINATION: Yes
Both eyes of all animals assigned to the study were examined just prior to the start of treatment, by means of both an ophthalmoscope and a slit-lamp microscope, after the instillation of 0.5% tropicamide (Visumidriatic, Merck, Sharp and Dohme, Rome, ltaly). The eyes of all animals in all groups were re-examined during week
13 of treatment. During the examination particular attention was paid to: anterior chamber, conjunctivae and eyelids, cornea and sclera, iris, lens, posterior segment - vitreous humour, and ocular fundus.

HAEMATOLOGY: Yes
- Time schedule for collection of blood: During week 13 samples of blood were withdrawn under light ether anaesthesia from the retro-orbital sinus of all rats from each group after overnight fasting.
- Anaesthetic used for blood collection: Yes (light ether)
- Animals fasted: Yes
- How many animals: all
- Parameters checked: haematocrit, haemoglobin, red blood cell count, reticulocyte count (not carried out as there were no signs of anaemia), mean red blood cell volume, mean corpuscular haemoglobin, mean corpuscular haemoglobin concentration, white blood cell count, differential leucocyte count (neutrophils, lymphocytes, eosinophils, basophils, monocytes, large unstained cells), abnormalities of the blood film, platelets, and prothrombin time

CLINICAL CHEMISTRY: Yes
- Time schedule for collection of blood: During week 13 samples of blood were withdrawn under light ether anaesthesia from the retro-orbital sinus of all rats after overnight fasting.
- Animals fasted: Yes
- How many animals: all
- Parameters checked: alkaline phosphatase, alanine aminotransferase, aspartate aminotransferase, urea, creatinine, glucose, albumin, total bilirubin, total cholesterol, total protein, sodium, phosphorus, potassium, calcium, and chloride.

URINALYSIS: Yes
- Time schedule for collection of urine: During week 13 samples of overnight urine samples were collected from all rats
- Metabolism cages used for collection of urine: Not specified
- Animals fasted: Yes, collection of urine during overnight fasting
- Parameters checked: appearance, volume, specific gravity, pH, protein, total reducing substances, glucose, ketones, bilirubin, urobilinogen, blood, and sediment.

NEUROBEHAVIOURAL EXAMINATION: Yes
- Time schedule for examinations: at the end of the study
- Dose groups that were examined: all
- Battery of functions tested: sensory activity / grip strength / motor activity / other:
The Motor Activity of the first 5 males and 5 females was measured once during week 12 of treatment by an automated activity recording device (Mini Opto-V arimex, Columbus International Corp.). Measurements were performed using a computer generated random order.

IMMUNOLOGY: No

Sacrifice and pathology:

GROSS PATHOLOGY: Yes

HISTOPATHOLOGY: Yes, broad range of organs

Organ weights: adrenal glands, brain, epididymides, heart, liver, kidneys, ovaries, spleen, testes, thymus, and uterus.

Statistics:

Standard deviations were calculated as considered appropriate. For continuous variables the significance of the differences amongst groups was assessed by analysis of variance. Differences between each treated group and the control group were assessed by Dunnett's test using a pooled error variance. The homogeneity of the data was verified by Bartlett's test. If data were found to be inhomogeneous a Modified t test (Cochran and Cox) was applied. The mean values, standard deviations and statistical analysis were calculated from the actual values in the computer without rounding off. Microscopic observations were tested for statistical significance using the non-parametric Kolmogorov-Smirnov test.

Results and discussion

Results of examinations

Clinical signs:

effects observed, treatment-related

Description (incidence and severity):

Salivation, mainly slight in severity, was observed in some treated as well as control animals during the study. The incidence of this sign was higher in treated males than in controls while the incidence in treated female groups was almost comparable to control animals. As this sign was noted in both control and treated animals it could be considered to be due to the vehicle used for the administration of the test item.
At the daily pre- and post-dose observations, salivation was occasionally noted just before dosing or 30 minutes after dosing, in a few male and female animals from the low, mid- and high-dose groups.

Mortality:

no mortality observed

Body weight and weight changes:

effects observed, non-treatment-related

Description (incidence and severity):

No changes in body weight were observed in males and females from mid- and high-dose groups and females from the low-dose group. However, a reduced body weight increment was observed in low-dose males when compared to controls, mainly from the 3rd to the 6th week of dosing, causing a change in body weight, which achieved statistically significance in most ofthe measurements performed from the 4th week to the end of treatment.
This change, noted in only one sex receiving the low dose-level, was not considered treatment-related.
Terminal body weight was significantly reduced in low-dose males.

Food consumption and compound intake (if feeding study):

effects observed, non-treatment-related

Description (incidence and severity):

Reductions in food consumption of low-dose males were observed on 4 occasions during the study (weeks 2, 3, 5 and 10).
Low-dose females and mid- and high-dose animals did not show any changes in food consumption when compared to controls throughout the study.

Food efficiency:

not examined

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

not examined

Ophthalmological findings:

no effects observed

Haematological findings:

no effects observed

Clinical biochemistry findings:

no effects observed

Urinalysis findings:

no effects observed

Behaviour (functional findings):

no effects observed

Immunological findings:

not examined

Organ weight findings including organ / body weight ratios:

no effects observed

Gross pathological findings:

effects observed, non-treatment-related

Description (incidence and severity):

No relevant macroscopic changes were detected in the animals killed at termination which could be considered related to the test item. The macroscopic observations reported did not reveal any significant difference between treatment and control animals killed at termination.
Statistically significant reductions in absolute liver and kidney weights were achieved in low and high-dose males andin relative weights in mid and high-dose males while the absolute liver weight was increased in low-dose females. In addition, absolute spleen weight was increased in mid- and high-dose females and absolute and relative uterus weight was decreased in low-dose females. All these changes were considered to be of no biological relevance as they were not consistent between sexes and not related to any morphological changes at the microscopic examination.

Neuropathological findings:

no effects observed

Histopathological findings: non-neoplastic:

effects observed, non-treatment-related

Description (incidence and severity):

The histopathological changes detected did not show any evident differences between the treated and control animals that could be considered treatment-related.
The seminiferous tubules in the testes were evaluated with respect to their stage in the spermatogenic cycle and the integrity of the various cell type present within the different stages. The findings observed were considered to be an expression of spontaneous pathology normally seen in this species.

Histopathological findings: neoplastic:

no effects observed

Details on results:

No abnormalities were detected at the macroscopic observation and histopathological examination of the organs of the reproductive systems such as prostate gland, seminal vesicles, testes and epididymides in males and ovaries and uterus in females. The weights of testes, epididymides, ovaries and uterus were not affected by treatment. In addition, following the evaluation of the seminiferous tubules with respect to their stage in the spermatogenic cycle and the integrity of the various cell type present within the different stages, no abnormalities were noted.

Effect levels

Key result

Dose descriptor:

NOAEL

Effect level:

1 000 mg/kg bw/day (nominal)

Based on:

test mat.

Remarks:

highest dose tested

Sex:

male/female

Basis for effect level:

other: absence of adverse toxic effects

Target system / organ toxicity

Key result

Critical effects observed:

no