Merginol 901

Repeated dose toxicity: oral

Administrative data

Endpoint:

repeated dose toxicity: oral, other

Type of information:

read-across based on grouping of substances (category approach)

Adequacy of study:

key study

Justification for type of information:

Please refer to chapter 13 for the category justification.

Cross-reference

Cross-reference 1

Reason / purpose for cross-reference:

read-across: supporting information

Reference

Endpoint:

sub-chronic toxicity: oral

Type of information:

experimental study

Adequacy of study:

supporting study

Study period:

1990-06-18 until 1990-09-21

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

comparable to guideline study with acceptable restrictions

Qualifier:

equivalent or similar to guideline

Guideline:

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

Deviations:

yes

Remarks:

Application was performed on 5 days per week as opposed to 7 days per week with no analytical verification of test concentrations.

GLP compliance:

yes

Limit test:

no

Specific details on test material used for the study:

- Name of test material: Stabiol VP 1711
- Substance type: fatty acid ester
- Physical state: white powder
- Lot/batch No.: 041/8/055 (1988-02-04)
- Stability under test conditions: stable under test conditions

Species:

rat

Strain:

Sprague-Dawley

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Charles River Laboratories, Sulzfeld, Germany
- Age at study initiation: 4 weeks
- Weight at study initiation: 62 - 82 g (females); 58 - 80 g (males)
- Housing: Makrolon Cage type III
- Diet: Altromin 1324, ad libitum, supplied by Altromin GmbH
- Water: tap water (drinking water quality analytically verified), ad libitum
- Acclimation period: 13 days

Route of administration:

oral: gavage

Vehicle:

peanut oil

Details on oral exposure:

PREPARATION OF DOSING SOLUTIONS:
The testing solution was prepared daily just before administration by weighing out an appropriate amount of test substance and dispersing it in the vehicle.

VEHICLE
- The administration volume was 5 mL/kg bw.
- The concentration of test item in the vehicle was: 200, 60, 20 and 0 mg/mL

Analytical verification of doses or concentrations:

not specified

Details on analytical verification of doses or concentrations:

No data

Duration of treatment / exposure:

107 to 108 days starting with animal acclimation, with a total treatment of 68 to 69 administrations

Frequency of treatment:

5 days per week

Dose / conc.:

100 mg/kg bw/day (actual dose received)

Dose / conc.:

300 mg/kg bw/day (actual dose received)

Dose / conc.:

1 000 mg/kg bw/day (actual dose received)

No. of animals per sex per dose:

10 animals/ sex/ dose.
Satellite groups of 5 male and 5 female animals were used for the control and 1000 mg/kg bw day dose groups.

Control animals:

yes, concurrent vehicle

Details on study design:

- On the day of arrival the animals were subjected to an acclimatization period during which they received ground diet and drinking water ad libitum.
- The total number of application was 68 to 69, corresponding to a total administered amount of 6800 to 6900, 20400 to 20700, 68000 to 69000 mg/kg bw for the respective 100, 300, 1000 mg/kg bw/day dose groups. -
- Satellite groups of 5 male and 5 female animals were used for the control and 1000 mg/kg bw day dose groups. Following concurrent treatment these satellite animals were not treated for an additional observation period of 33 days in order to investigate the reversibility of potential toxic responses.

Positive control:

Not required

Observations and examinations performed and frequency:

MORTALITY
A check for moribund and dead animals was made twice daily on working days. If animals were in a moribund state, they were sacrificed and necropsied.

CLINICAL OBSERVATIONS
All animals were checked twice daily for any abnormal clinically signs on working days.

FOOD CONSUMPTION
Group food consumption was determined weekly for each cage.

DRINKING WATER CONSUMPTION
Drinking water consumption was determined weekly for each cage.

BODY WEIGHT
Body weight was determined before the start of the administration period. During the administration period body weight was determined on study day 0 (start of the administration period) and thereafter at weekly intervals.

HAEMATOLOGY
The following parameters were determined in blood: Leukocyte count (WBC), Erythrocyte count (RBC), Hemoglobin (HGB), Hematocrit (HCT), Mean corpuscular volume (MCV), Platelet count (PLT), Differential blood count

CLINICAL CHEMISTRY
Alanine aminotransferase (ALT), Aspartate aminotransferase (AST), Alkaline phosphatase (AP), γ-Glutamyltransferase (GGT), Sodium (NA), Potassium (K), Chloride (CL), Calcium (CA), Urea (UREA), Creatinine (CREA), Glucose (GLUC), Total bilirubin (TBIL), Total protein (TPROT), Cholesterol (CHOL)

OPHTHALMOSCOPY
Prior to the start of necropsy the eyes of all animals of the control and high dose animals were treated with Mydriaticum and examined for any changes using a slit lamp.

Sacrifice and pathology:

NECROPSY
The animals were sacrificed by exsanguination under an overdose of ether. The exsanguinated
animals were necropsied and assessed by gross pathology.

ORGAN WEIGHTS
The following weights were determined in all animals sacrificed on schedule: Anesthetized animals, Adrenal glands, Brain, Heart, Kidneys, Liver, Spleen, Testes, Thymus

HISTOPATHOLOGY
Fixation was followed by histotechnical processing, examination by light microscopy and assessment of findings on following organs: Adrenal glands, Aorta, Skeletal muscle, Brain, Cecum, Uterus, Coagulating glands, Colon, Duodenum, Epididymis (left), Esophagus, Heart, Ileum, Jejunum, Kidneys, Liver, Lung, Lymph nodes (mesenteric and axillary lymph nodes), Pancreas, Tounge, Pituitary gland, Prostate, Salivary gland, Seminal vesicles, Skin, Semen vesicles, Spleen, Forestomach, Testis, Thymus, Thyroid glands, Trachea, Uterus

Statistics:

- A comparison of inter-group differences (clinical chemistry, hematology and body weight) was performed using the t-test.
- DUNNETT's test was used to test the hypothesis of equal means.
- Steel-test was used to evaluate the inter-group differences relating to organ weights of each dose group.

Clinical signs:

no effects observed

Description (incidence and severity):

No test substance-related findings were observed

Mortality:

no mortality observed

Description (incidence):

In the present study no animal died ahead of schedule.

Body weight and weight changes:

no effects observed

Description (incidence and severity):

No test substance-related findings were observed in all parameters under investigation.

Food consumption and compound intake (if feeding study):

no effects observed

Description (incidence and severity):

No test substance-related findings were observed.

Food efficiency:

no effects observed

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

no effects observed

Description (incidence and severity):

No test substance-related findings were observed.

Ophthalmological findings:

no effects observed

Description (incidence and severity):

No test substance-related findings were observed in all parameters under investigation.

Haematological findings:

effects observed, non-treatment-related

Description (incidence and severity):

A decrease of the haematocrit (HCT) value in the male groups 2 - 4 was observed. The decrease of the HCT seems to be compound-and partially dose-related. Additionally the intermediate analysis showed slightly reduced Red Blood Cell (RBC)-value for the male group 4. The observed deviation of the HCT is considered to be incidental/ because the diagnosed values are within the ranges of the historical control. In addition to this, there are no corresponding deviations of the RBC- or MCV-values to prove the biological relevance of this findings.

Clinical biochemistry findings:

no effects observed

Description (incidence and severity):

No test substance-related findings were observed in all parameters under investigation.

Urinalysis findings:

not examined

Behaviour (functional findings):

no effects observed

Immunological findings:

not examined

Organ weight findings including organ / body weight ratios:

no effects observed

Description (incidence and severity):

- Absolute organ weights: No test substance-related findings were observed in all parameters under investigation.
- Relative organ weights: No test substance-related findings were observed in all parameters under investigation.

Gross pathological findings:

no effects observed

Description (incidence and severity):

The macroscopical examination of the organs displayed some observations like discolouration of the thymus deformation of the spleen, cyst of the kidney, hydronephrosis, atrophy of the testes, hydrometra and necrosis of the fatty tissue which were considered to be spontaneous. Compound-related macroscopical effects were not observed. The male and female animals of the recovery group 4 (recovery period 33 days) showed no macroscopical compound-related alterations.

Neuropathological findings:

not examined

Histopathological findings: non-neoplastic:

effects observed, non-treatment-related

Description (incidence and severity):

No test substance-related findings were observed in all parameters under investigation. However, in the male and female animals of all groups (including the recovery group 1 and 4) the livers, the heart and the mandibulary lymph node showed effects which were due to a bacteriosis of unknown etiology. The observed germinal hyperplasia of the mandibulary lymph node can be interpreted as a consequence of this bacteriosis.

Histopathological findings: neoplastic:

no effects observed

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 at the highest dose tested.

Critical effects observed:

no

Cross-reference 2

Reason / purpose for cross-reference:

read-across: supporting information

Reference

Endpoint:

short-term repeated dose toxicity: oral

Type of information:

experimental study

Adequacy of study:

supporting study

Study period:

1985-09-23 until 1985-11-22

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

comparable to guideline study with acceptable restrictions

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 407 (Repeated Dose 28-Day Oral Toxicity Study in Rodents)

Deviations:

yes

Remarks:

Application was performed on 5 days per week as opposed to 7 days per week

GLP compliance:

yes

Limit test:

no

Specific details on test material used for the study:

- Name of test material: LA 990
- Substance type: organic
- Physical state: viscous liquid, beige-coloured
- Lot/batch No.: 37-4-222
- Stability under test conditions: stable under test conditions
- Storage condition of test material: at room temperature

Species:

rat

Strain:

Sprague-Dawley

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Charles River Laboratories, Sulzfeld, Germany
- Age at study initiation: 4 weeks
- Weight at study initiation: 60- 81 g (females); 59 - 78 g (males)
- Housing: Makrolon Cage type III
- Diet: Altromin 1324 - ad libitum- supplied by Altromin GmbH
- Water: tap water (drinking water quality analytically verified) - ad libitum
- Acclimation period: 10 days

Route of administration:

oral: gavage

Vehicle:

other: 1% Carboxymethylcellulose and 0.5% Cremophor (CMCC)

Details on oral exposure:

PREPARATION OF DOSING SOLUTIONS:
The testing suspension was prepared by weighing out an appropriate amount of test substance and dispersing it in the vehicle.

VEHICLE
- The test item was applied as a suspension in CMCC (vehicle).
- The administration volume was 10 mL/kg body weight.
- The concentration of test item in the vehicle was: 100, 50, 10 and 0 mg/mL.

Analytical verification of doses or concentrations:

not specified

Details on analytical verification of doses or concentrations:

No data

Duration of treatment / exposure:

42 to 43 days starting with animal acclimation, with a total treatment of 23 to 24 administrations

Frequency of treatment:

5 days per week

Dose / conc.:

100 mg/kg bw/day (actual dose received)

Dose / conc.:

500 mg/kg bw/day (actual dose received)

Dose / conc.:

1 000 mg/kg bw/day (actual dose received)

No. of animals per sex per dose:

10 animals
Satellite groups of 5 male and 5 female animals were used for the control and 1000 mg/kg bw day dose groups. Following concurrent treatment these satellite animals were not treated for an additional observation period of 28 days in order to investigate the reversibility of potential toxic responses

Control animals:

yes, concurrent vehicle

Details on study design:

- On the day of arrival the animals were subjected to an acclimation period during which they received ground diet and drinking water ad libitum.
- The total number of application was 23 to 24, corresponding to a total administerd amount of 2300 to 2400, 11500 to 12000, 23000 to 24000 mg/kg bw for the respective 100, 500, 1000 mg/kg bw/day dose groups.
- Satellite groups of 5 male and 5 female animals were used for the control and 1000 mg/kg bw day dose groups. Following concurrent treatment these satellite animals were not treated for an additional observation period of 28 days in order to investigate the reversibility of potential toxic responses

Positive control:

Not required

Observations and examinations performed and frequency:

MORTALITY
A check for moribund and dead animals was made twice daily on working days. If animals were in a moribund state, they were sacrificed and necropsied.

CLINICAL OBSERVATIONS
All animals were checked twice daily for any abnormal clinically signs on working days.

FOOD CONSUMPTION
Group food consumption was determined weekly for each cage.

DRINKING WATER CONSUMPTION
Drinking water consumption was determined weekly for each cage.

BODY WEIGHT
Body weight was determined before the start of the administration period. During the administration period body weight was determined on study day 0 (start of the administration period) and thereafter at weekly intervals.

HAEMATOLOGY
The following parameters were determined in blood: Leukocyte count (WBC), Erythrocyte count (RBC), Hemoglobin (HGB), Hematocrit (HCT), Mean corpuscular volume (MCV), Platelet count (PLT), Differential blood count,

CLINICAL CHEMISTRY
Alanine aminotransferase (ALT), Aspartate aminotransferase (AST), Alkaline phosphatase (AP), γ-Glutamyltransferase (GGT), Sodium (NA), Potassium (K), Chloride (CL), Calcium (CA), Urea (UREA), Creatinine (CREA), Glucose (GLUC), Total bilirubin (TBIL), Total protein (TPROT), Cholesterol (CHOL)

OPHTHALMOSCOPY
Prior to the start of necropsy the eyes of all animals of the control and high dose animals were treatd with Mydriaticum and examined for any changes using a slit lamp.

Sacrifice and pathology:

NECROPSY
The animals were sacrificed by exsanguination under an overdose ether. The exsanguinated animals were necropsied and assessed by gross pathology.

ORGAN WEIGHTS
The following weights were determined in all animals sacrificed on schedule: Anesthetized animals, Adrenal glands, Brain, Heart, Kidneys, Liver, Spleen, Testes, Thymus

HISTOPATHOLOGY
Fixation was followed by histotechnical processing, examination by light microscopy and assessment of findings on following organs: Adrenal glands, Aorta, Skeletal muscle, Brain, Cecum, Uterus, Coagulating glands, Colon, Duodenum, Epididymis (left), Esophagus, Heart, Ileum, Jejunum, Kidneys, Liver, Lung, Lymph nodes (mesenteric and axillary lymph nodes), Pancreas, Tounge, Pituitary gland, Prostate, Salivary gland, Seminal vesicles, Skin, Semen vesicles, Spleen, Forestomach, Testis, Thymus, Thyroid glands, Trachea, Uterus

Statistics:

- A comparison of inter-group differences (clinical chemistry, hematology and body weight) was performed using the t-test.
- Steel-test was used to evaluate the inter-group differences relating to organ and bone weights of each dose group.

Clinical signs:

no effects observed

Description (incidence and severity):

No test substance-related findings were observed

Mortality:

no mortality observed

Description (incidence):

In the present study no animal died ahead of schedule.

Body weight and weight changes:

no effects observed

Description (incidence and severity):

No test substance-related findings were observed in all parameters under investigation.

Food consumption and compound intake (if feeding study):

effects observed, treatment-related

Description (incidence and severity):

Salivation was observed following substance administration - No further test substance-related findings were observed.

Food efficiency:

not examined

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

no effects observed

Description (incidence and severity):

No test substance-related findings were observed.

Ophthalmological findings:

not examined

Haematological findings:

effects observed, treatment-related

Description (incidence and severity):

A decrease of the haematocrit (HCT) value and Red Blood Cell (RBC) were observed for male animals in the high dose group. In female animals the same findings occurred in the middle dose group.

Clinical biochemistry findings:

effects observed, treatment-related

Description (incidence and severity):

A slight significant increase in GPT-levels in males of the high dose and a significant increase of GPT-values in female animals of the high dose was measured.

Urinalysis findings:

not examined

Behaviour (functional findings):

not examined

Immunological findings:

not examined

Organ weight findings including organ / body weight ratios:

effects observed, treatment-related

Description (incidence and severity):

The relative liver weights were significantly increased in male animals of the high dose. A slight increase of the relative liver weights was seen in female animals of the high dose. These findings were considered to be compound related.

Gross pathological findings:

no effects observed

Description (incidence and severity):

The macroscopical examination of the organs did not reveal any compound related changes.

Neuropathological findings:

not examined

Histopathological findings: non-neoplastic:

effects observed, treatment-related

Description (incidence and severity):

Microscopic examination in animals of high dose level revealed degenerative alterations in kidneys which were considered to be reversible in the recovery group.

Histopathological findings: neoplastic:

no effects observed

Dose descriptor:

NOAEL

Effect level:

100 mg/kg bw/day (actual dose received)

Based on:

test mat.

Sex:

male/female

Basis for effect level:

histopathology: non-neoplastic

organ weights and organ / body weight ratios

Remarks on result:

other: see any other information on results incl tables

Critical effects observed:

no

It is argueable whether the effects observed in the study are sufficient to conclude on a NOAEL of 100 mg/kg bw/d. According to a recent publication by Hall et al., the variation of liver weights without histological evidence or distinct alterations in clinical chemistry is not considered to be an adverse effect. Hall et al. reach the conclusion that only a 2-3-fold increase in ALT (GPT) or a biologically significant change in other biomarkers is considered to be an adverse effect. In the study at hand, the only remarkable alteration in liver parameters was an ALT increase in the highest dose group of males and females, but it was less than 2-fold. Regarding the observed degradations in the kidney, they were not accompanied with relevant biochemical changes (urea, creatinie, sodium, potassium) and were reversible in the recovery group. The absence of adverse effects in the 90-day study further supports the argumentation that the alterations found in the 28-day study were temporary and rather a non-adverse adaptation than an evidence for a toxicological action.

Reference

Hall et al. (2012), Liver hypertrophy: a review of adaptive (adverse and non-adverse) changes – conclusions from the third international ESTP workshop, Toxicol Pathol, 40(7), 971-94

Materials and methods

Test material

Test material information

Constituent 1

Reference substance name:

Fatty acids, C16-18 and C18-unsatd., epoxidized, Me esters, oligomeric reaction products with trimethylolpropane

Molecular formula:

Not applicable (UVCB substance)

IUPAC Name:

Fatty acids, C16-18 and C18-unsatd., epoxidized, Me esters, oligomeric reaction products with trimethylolpropane

Results and discussion

Effect levels

1

Key result

Dose descriptor:

NOAEL

Remarks:

90 days

Effect level:

1 000 mg/kg bw/day (actual dose received)

Based on:

test mat.

Sex:

male/female

Basis for effect level:

other: No toxic effects at highest dose tested

Remarks on result:

other: Read-across from 151661-88-0, sub-chronic

2

Dose descriptor:

NOAEL

Remarks:

28 days

Effect level:

100 mg/kg bw/day (actual dose received)

Based on:

test mat.

Sex:

male/female

Basis for effect level:

histopathology: non-neoplastic

organ weights and organ / body weight ratios

Remarks on result:

other: Read-across from Oleic methyl ester, epoxidized, reaction products with glycerol (OEG), sub-acute

Target system / organ toxicity

Key result

Critical effects observed:

no