Fatty acids, tall-oil, compds. with triethanolamine

Repeated dose toxicity: oral

Administrative data

Endpoint:

sub-chronic toxicity: oral

Type of information:

experimental study

Adequacy of study:

key study

Study period:

6th August 1974 to 8 December 1977

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

comparable to guideline study with acceptable restrictions

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

1977

Report date:

1977

Materials and methods

Test guideline

Qualifier:

equivalent or similar to guideline

Guideline:

other: Two generation Reproduction Study in rat

GLP compliance:

no

Remarks:

Study pre-dates GLP

Limit test:

no

Test material

Test material information

Constituent 1

Reference substance name:

2,2',2''-nitrilotriethanol

EC Number:

203-049-8

EC Name:

2,2',2''-nitrilotriethanol

Cas Number:

102-71-6

Molecular formula:

C6H15NO3

IUPAC Name:

2,2',2''-nitrilotriethanol

Test material form:

not specified

Test animals

Species:

rat

Strain:

Sprague-Dawley

Sex:

male

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Age at study initiation: 80 days of age
- Housing - All rats were group housed in suspended stainless steel cages, five per cage
-Diet - ad libitum
Water - ad libitum

Administration / exposure

Route of administration:

oral: feed

Vehicle:

unchanged (no vehicle)

Details on oral exposure:

DIET PREPARATION
-Rate of prepartation of diet: TEsts diets were prepared weekly by mixing the correct amount of the respective compounds with the appropriate amount of basic diet in a three cubic foot Patterson Kelley twin shell mixer, tumbling at a rate of 40 tumbles per minute around a high speed coaxial mixing bar equipped with discs bearing slanted blades and rotating at the rate of 2,000 RPM
-The tests diets were prepared weekly by the supervisor on a percent weight of diet basis. Half of the control feed was first poured into the mixture, followed by the test material or test matrial containing higher diet, followed by the other hald of the control diet. The mixer was spun for 15 minutes for each diet level.
-Feed was offered ad libitum. All feed remaining at the end of the week was destroyed

Analytical verification of doses or concentrations:

no

Duration of treatment / exposure:

P0 animals were treated for 20 days prior to mating the during mating and weaning
F1 animals were weaned onto test diet and fed it until mating at 100 days of age and throughout mating and weaning

Frequency of treatment:

Continuous in the diet

Doses / concentrations

Doses / concentrations 1

Dose / conc.:

5 other: %

Remarks:

4500 (female) and 4000 (male) mg/kg/day (5% equates to 50000 mg.kg diet, multiplied by 0.09 for females and 0.08 for males using the EFSA guidance to give a dose of 4500 or 4000 mg/kg for females and males, respectively

Doses / concentrations 2

Dose / conc.:

10 other: %

Remarks:

9000 (female) and 8000 (male) mg/kg/day (10% equates to 1000,000 mg/kg diet, multiplied by 0.09 for females and 0.08 for males using the EFSA guidance.

No. of animals per sex per dose:

P0 - 15 males/dose, 30 females/dose
F1 - 20 animals/per sex/dose

Control animals:

yes, plain diet

Examinations

Observations and examinations performed and frequency:

CAGE SIDE OBSERVATIONS: Yes

- All rats were inspected by the animal caretaker daily and by the supervising technician once a week.
-Any abnormal signs observed by the animal caretaker were brought to the supervising technician's attention immediately, who followed it routinely either to clinical recovery or moribund condition whereupon the animal was sacrificed. Pertinent signs were called to the directors attention

Sacrifice and pathology:

ORGAN WEIGHTS
- The following organs were weighed from ten male and ten female rats from each group in the F1 generation: Thyroids; Heart; Liver; Adrenals; Kidneys; and Gonads. These organs were weighed after fixation in 10% buffered formalin.

NECROPSY
- All rats, whether they died or were sacrificed, received a complete gross necropsy (F1 and F2 generations).
- Rats were fasted overnight prior to necropsy. All rats except those designated for clinical chemistry and organ weights were euthanised in an ether jar. The rats intended for clinical chemistry and organ weights were anesthetised with ether, their abdomen opened and their blood drawn from the inferior vena cava behind and below the liver.
- Each rat was subsequently pinned on a board with its extremities extended and their ventral skin reflected over their head exposing the thoracic, abdominal and pelvic cavities. The viscera were examined in situ. The neck of the animal was subsequently severed in such a way as to leave the head attached to the trachea and oesophagus, the viscera removed in toto and the total tissue aggregate immersed in buffered 10% formalin (W/V ratio: approx.: 1:10). Representative blocks of skin, muscle, bone (lumbar vertebra) and peripheral (sciatic) nerve were also immersed.
- Organ weighing and tissue blocking were done after fixation for at least 72 hours. Blocking was done in double labelled plastic cassettes. The following tissues were examined histologically from each F1 rat used in the determinations listed above as well as on ten male and ten female F2 weanlings per group with the remainder destroyed: Brain; Pituitary gland; Spinal Cord; Eye; Sub-maxillary gland; Thyroids; Lungs; Heart; Liver; Kidneys; Spleen; Adrenals; Pancreas; Stomach; Intestines; Lymph Nodes; Bladder; Gonads; Skin; Bone and marrow; Nerve and Muscle; and Any unusual lesions.
Only those animals showing significant microscopic findings are submitted herewith on Individual Animal Data Records (I.A.D.Rs).

HISTOLOGY PROCEDURES
- Once the animal tissues were blocked, they were sent to histology. The cassettes were then processed by a three phased automated procedure extending over a 16 hour period and involving the following steps: Dehydration of tissues; Clearing of tissues by xylene and Infiltration of tissue with paraffin. Bone tissue was decalcified prior to processing by placing it into a solution of hydrochloric acid and chelating agent for a period of time. Embedding was performed using a Tissue Tek II Tissue Embedding Centre. It involved placing tissue into stainless steel melds, spacially arranging tissue within the mould so that no two pieces were tangent, filling mould with paraffin and placing the same plastic cassette lid as used in processing onto the mould. Now referred to as "blocks" the tissue specimens were cooled, solidified and finally removed from the moulds.
- Paraffin blocks were cut on a Spencer 820 American Optical microtome at a thickness of five micra. This thin tissue section was subsequently transferred to a clean glass slide with an identification number on it. Slides were routinely stained with haematoxylin and eosin. The actual staining procedure required several additional solutions to give acceptable results. To insure good quality control, the histologist carefully monitored these solutions and maintained same fresh and non-contaminated. A microscopic check was made for cellular detail and differentiation prior to for-warding the slides to the pathologist. Upon request certain slides received a "special stain" for diagnostic purposes. In these instances the histologist ran a positive control slide concurrently to confirm that the stain was acceptable.

Results and discussion

Results of examinations

Clinical signs:

no effects observed

Mortality:

not examined

Body weight and weight changes:

not examined

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:

not examined

Urinalysis findings:

not examined

Behaviour (functional findings):

not examined

Immunological findings:

not examined

Organ weight findings including organ / body weight ratios:

not examined

Gross pathological findings:

not examined

Neuropathological findings:

not examined

Histopathological findings: non-neoplastic:

not examined

Histopathological findings: neoplastic:

not examined

Other effects:

not examined

Effect levels

1

Dose descriptor:

NOAEL

Effect level:

9 000 mg/kg bw/day (nominal)

Based on:

test mat.

Sex:

female

Basis for effect level:

other: No test material related effects observed

2

Dose descriptor:

NOAEL

Effect level:

8 000 mg/kg bw/day (nominal)

Based on:

test mat.

Sex:

male

Basis for effect level:

other: No test material related effects observed

Target system / organ toxicity

Critical effects observed:

no

Applicant's summary and conclusion

Conclusions:

No test material related effects observed up to concentrations of 8000 and 9000 mg/kg for males and females, respectively.

Executive summary:

The reproductive toxicity of the test material was investigated in a 2-generation reproductive toxicity study with Sprague-Dawley rats.

During the study fifteen male and thirty female rats per group at 80 days of age were placed on diet at test material concentrations of 5 and 10%, alongside a negative control plain diet and a negative control of oleic acid at 5 and 10%. All animals were mated within each group at 100 days of age (F0 generation). The offspring were weaned onto the corresponding diet. Twenty male and 20 female rats from each group (with the members of each sex coming from a different dam) were selected shortly after weaning and were carried to sexual maturity. The F0 parents and all remaining offspring were subsequently destroyed. At 100 days of age the F1 animals were mated and parameters were recorded.

The test material doses received via the 5 % diet were calculated to be 4500 (female) and 4000 (male) mg/kg/day (5% equates to 50,000 mg/kg diet, multiplied by 0.09 for females and 0.08 for males using the EFSA guidance to give a dose of 4,500 or 4000 mg/kg/day, respectively) and at 10% diet were 9000 (female) and 8000 (male) mg/kg/day.

There were no test material related findings observed in any of the measured parameters or indices determined in the study.

Under the conditions of the study, the NOAEL for reproductive toxicity was determined to be 9000 mg/kg/day for females and 8000 mg/kg/day for males.

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