Corn oil (Zea mays, Gramineae), peroxidised

Repeated dose toxicity: oral

Administrative data

Endpoint:

short-term repeated dose toxicity: oral

Type of information:

experimental study

Adequacy of study:

key study

Study period:

from 15 April 2011 to XXX

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: Study performed according to OECD guideline and GLP

Cross-reference

Reason / purpose for cross-reference:

reference to same study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2011

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

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

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Limit test:

no

Test material

Test material information

Constituent 1

Reference substance name:

corn oil, peroxidised

IUPAC Name:

corn oil, peroxidised

Test animals

Species:

rat

Strain:

Wistar

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Velaz Prague, Czech Republic
- Age at study initiation: age of animals at study initiation was about 12 weeks
- Weight at study initiation: males 214.0g, females 169.2g (average body weights at the start of the study)
- Fasting period before study: no
- Housing: in cages (number of animals in cage according period of the study) in a room equipped with central air-conditioning
- Diet: certified laboratory food (MP-OŠ-06 extrudes, Snina, Slovak Republic) was offered in recommended doses
- Water (e.g. ad libitum): tap water for human consumption (ad libitum)
- Acclimation period: acclimatized to the environmental conditions for 5 days prior to the start of the study

ENVIRONMENTAL CONDITIONS
- Temperature: 22 ± 2° C
- Humidity: 55 ± 10%
- Air changes: no data
- Photoperiod: 12-hour light /12-hour dark cycle

IN-LIFE DATES: From 18 April 2011 to 31 October 2011

Administration / exposure

Route of administration:

oral: gavage

Vehicle:

olive oil

Details on oral exposure:

PREPARATION OF DOSING SOLUTIONS: no data

VEHICLE
- Justification for use and choice of vehicle:
- Concentration in vehicle:
- Amount of vehicle (if gavage): dose volume was 5 ml/kg of body weight and afterwards was adjusted individually according to the weight development of the animals
- Lot/batch no. (if required): 100 (Olificio Lucca, Italy)

Analytical verification of doses or concentrations:

no

Details on analytical verification of doses or concentrations:

Analytical verification was not possible, as the product (an oil) was be mixed with an other oil (not possible to separate by analysis the components of the test substance from those from the vehicle, as both are oils containing similar fatty acids).

Duration of treatment / exposure:

The all animals were treated with the test article
- 14 days pre-mating,
- 14 days mating (maximum)
- 22 days gestation (approximately)
- 4 days lactation

Frequency of treatment:

Every day (about 9:00 am)

Doses / concentrations

Remarks:

Doses / Concentrations:
0, 100, 300, 1000
Basis:
other: mg substance/kg bw

No. of animals per sex per dose:

12 animals per sex per dose
+ 6 animals per sex per dose in the satellite groups at 0 and 1000 mg/kg bw (recovery groups)

Control animals:

yes, concurrent vehicle

Details on study design:

- Dose selection rationale: doses of test article were determined according to results of Dose Range Finding Assay
- Rationale for selecting satellite groups: control and high dose
- Post-exposure recovery period in satellite groups: 14 days

Positive control:

none

Examinations

Observations and examinations performed and frequency:

CAGE SIDE OBSERVATIONS: Yes
- Time schedule: once a day, at the same time, considering the peak period of anticipated effects after dosing.

DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: prior to the first exposure and at once a week thereafter.

BODY WEIGHT: Yes
- Time schedule for examinations: on the first day of dosing, weekly thereafter, and at the study termination. During pregnancy, females were weighed on days 0, 7, 14 and 20, day 1 and day 4 post-partum. Body weight measurements of satellite animals (recovery period) were performed weekly.

FOOD CONSUMPTION: During pre-mating, pregnancy and lactation measurement of food consumption were made weekly. The measurements of food consumption during mating were made, but were not evaluated.

WATER CONSUMPTION: No

OPHTHALMOSCOPIC EXAMINATION: No

HAEMATOLOGY: Yes
- Time schedule for collection of blood: blood samples for haematology and clinical chemistry were collected before the first administration of the test article (0 sample) and at study termination (1st sample); in selected animals of the satellite groups after 14 day recovery period (2nd sample).
- Anaesthetic used for blood collection: No data
- Animals fasted: Yes
- How many animals: six males, six females per group and satellite animals
- Parameters checked: haematocrit, haemoglobin, leukocytes, erythrocytes, mean corpuscular volume, mean corpuscular haemoglobin, mean corpuscular haemoglobin concentration, platelets, differential count - lymphocytes, neutrophils, eosinophils, basophils and monocytes, prothrombin time, activated partial thromboplastin time.

CLINICAL CHEMISTRY: Yes
- Time schedule for collection of blood: blood samples for haematology and clinical chemistry were collected before the first administration of the test article (0 sample) and at study termination (1st sample); in selected animals of the satellite groups after 14 day recovery period (2nd sample).
- Animals fasted: Yes
- How many animals: six males, six females per group and satellite animals
- Parameters checked: Enzymes (alkaline phosphatase, aspartate aminotransferase, alanine aminotransferase), glucose, total cholesterol, triacylglycerols, creatinine, urea, total bilirubin, total proteins, albumin, calcium, inorganic phosphorus, natrium, potassium, chloride.

URINALYSIS: Yes
- Time schedule for collection of urine: during the last week of the study (in 6 males per group)
- Metabolism cages used for collection of urine: Yes
- Animals fasted: No data
- Parameters checked: specific gravity, leucocytes, nitrite (presence), pH, protein, glucose, ketones, urobilinogen, bilirubin, blood (erythrocytes).

NEUROBEHAVIOURAL EXAMINATION: No

Sacrifice and pathology:

GROSS PATHOLOGY: Yes
After completion of treatment all adult surviving animals which were not designated for posttreatment observation were sacrificed by application of an overdose of anaesthetic. All recovery animals were sacrificed after the end of the recovery period.
All adult animals were subjected to a full detailed gross necropsy. Examination of the outer surface of the body, all orifices, cranial cavity, thoracic cavity, gastric cavity and their contents were carried out. Special attention was paid to the organs of the reproductive system. The number of implantation sites was recorded. The counting of corpora lutea was recorded. Terminal body weights were measured in order to calculate relative organ weights.
The weights of following organs were determined in all males sacrificed at the end of treatment and after recovery period: epididymides, testes.
In addition, for six adult males and females randomly selected from each group, the following tissues were weighed: liver, kidneys, adrenals, thymus, spleen, brain, hearth.
Dead pups killed at day 4 post-partum or shortly, were examined externally for gross abnormalities.

HISTOPATHOLOGY: Yes
The following organs from six adult males and females, randomly selected from each group, were preserved and intended subsequent histopathological examination: all gross lesions, small and large intestines (including Peyer's patches), thyroid glands, thymus, trachea, lungs, heart, lymph nodes, brain, stomach, liver, spleen, kidneys, adrenals, urinary bladder, uterus, peripheral nerve, spinal cord.
Histopathological examinations were performed on organs and tissues of the selected animals from the control and high dose groups.

Statistics:

Individual results (the body weight, food consumption, haematology, clinical chemistry, relative weights of organs and reproduction parameters) obtained during the study were statistically evaluated using statistical programme Statgraphics. Statistical evaluation was conducted separately for males and females. Descriptive statistics (means and standard deviation) were performed for males and females in all dose groups. Non parametric Kruskal-Wallis test was applied. In the case of statistically significant results the Kruskal-Wallis test was followed by Mann-Whitney W test to determine which medians are significantly different from the one in control group. All statistical tests were carried out at 5% significance level.

Results and discussion

Results of examinations

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):

not examined

Ophthalmological findings:

not examined

Haematological findings:

no effects observed

Clinical biochemistry findings:

no effects observed

Urinalysis findings:

no effects observed

Behaviour (functional findings):

not examined

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:
There were no test article-related deaths of animals during the study. All animals lived through observation period without important visible signs of intoxication.
General clinical findings were typical for Wistar rats of this age. Neither change of health nor negative reactions were registered in satellite animals. The test article was very well tolerated by animals of all dose groups.

BODY WEIGHT AND WEIGHT GAIN:
No statistically significant differences in body weights between control and all dose groups of males were detected. In high dose females, (1000 mg.kg-1), average body weights were lower against controls after day 20 of pregnancy. This is related to the lower number of pups (or lower weight of litters) in this dose group. All remaining values of this parameter in females were identical with those of control group.

FOOD CONSUMPTION AND COMPOUND INTAKE:
The food consumption in males and females of all dose groups was similar to the control group, with no statistical significance.

HAEMATOLOGY:
All values of monitored parameters in both sexes during the study stayed within the normal range for this species. Changes in haematology parameters were not connected with treatment; they are results of interindividual and intraindividual variability for this species or have only statistical relevance. No test article related effect on the haematology and blood coagulation parameters were observed in this study.

CLINICAL CHEMISTRY:
At the end of treatment no differences in clinical chemistry parameters which can definitively be attributed to the administration of the test article were found. Average values at different time points in all test groups were within the normal range for most of parameters.
A dose relationship was not observed. Changes in clinical chemistry parameters were not connected with treatment; they are results of interindividual and intraindividual variability for this species or they have only statistical relevance or could be attributed to pregnancy (ALP, urea, total protein and albumin).
After recovery period no relevant differences could be ascertained in all clinical chemistry parameters.

URINALYSIS:
No significant changes against normal physiological conditions were detected. Small amounts of protein, ketones and presence of blood (erythrocytes) were observed occasionally. These findings present in urine are often considered as physiological for this species. Urine volume (collected during 6 hours) showed differences between control and dose groups but without dose relationship.

ORGAN WEIGHTS:
Relative weights of heart were increased significantly in males from low and high dose group in comparison to the control group. No similar finding was seen in females. Relative weights of kidney left were increased significantly in males from high dose group in comparison to control group, however, not in females. Relative liver weights of female rats from high and medium dose groups were decreased significantly in comparison to control. Historically, it is not uncommon to see the relative weight of liver increased in treated animals versus controls. Also, no significant differences were observed in males or between recovery groups (high dose group and control). However, the effect of treatment cannot be excluded.

GROSS PATHOLOGY:
All animals survived until the ending of the observation period. Occasional macroscopical lesions during necropsy of rats were observed. Hypoplasia of testes and solid dark mass in epididymis in one male (low dose) were found. Histopathological examination was confirmed by degenerative changes of the spermiogenesis. These findings could be due to the pre-existing condition. Detection of testicular hypoplasia is common in Wistar rats. The light colour of liver in two females of low dose and two females of medium dose was observed.

HISTOPATHOLOGY: NON-NEOPLASTIC
There were no pathological changes that could be related to the effect of Peroxidised Corn Oil.

Effect levels

Dose descriptor:

NOAEL

Effect level:

>= 1 000 mg/kg bw/day (nominal)

Based on:

test mat.

Sex:

male/female

Basis for effect level:

other: overall effects

Target system / organ toxicity

Critical effects observed:

not specified

Applicant's summary and conclusion

Conclusions:

The no observed adverse effect level for systemic toxicity in males and females was concluded to be 1000 mg/kg bw.

Executive summary:

In a subacute toxicity study (OECD 422, GLP) peroxidised corn oil (in olive oil as vehicle) was administered to 12 Wistar rats/sex/dose in by gavage at dose levels of 0 (olive oil), 100, 300 and 1000 mg/kg bw/day. In the control and high dose groups, satellite animals were included (6 per sex and group) to provide information about reversibility of toxicological findings.

Females were treated 14 days before mating, 14 days during mating, 22-24 days during gestation and 4 days during lactation. Males were treated 56 days.

There were no test article-related deaths of animals during the study. All animals lived through observation period without important visible signs of intoxication. Neither change of health nor negative reactions were registered in the satellite animals. The test article was very well tolerated by animals of all dose groups.

The body weight of all animals moderately increased. No significant differences were noted between body weight animals of the control and all dose groups during treatment and recovery period.

The food consumption in males and females of all dose groups was similar to the control group.

No test article related effects on the haematology and blood coagulation parameters were observed.

No differences in clinical chemistry parameters which could definitively be attributed to the administration of the test article were found. After recovery period, no relevant differences were found in clinical chemistry parameters.

No significant changes against normal physiological levels were detected in urine of males.

The significant decrease of liver relative weight found in females of medium and high dose groups versus control could be related to the effect of the test article. Historically, it is not uncommon to see the relative weight of liver increased in treated animals versus controls. Also, no significant differences were observed in males or between recovery groups (high dose group and control). However, the effect of treatment cannot be excluded. However, no similar changes were seen in males.

Peroxidised Corn Oil administered at doses up to 1000 mg/kg bw/day caused neither treatment-related deaths of animals nor visible treatment-related signs of intoxication. Test article did not cause pathological and histopathological changes in organs and tissues of rats.

 

The NOAEL for systemic toxicity in males and females was concluded to be 1000 mg/kg bw.

 

 This subacute toxicity study in the rats is acceptable and satisfies the guideline requirement for a Repeated Dose Toxicity Study (OECD 422) in rats.