6-nonyl-1,3,5-triazine-2,4-diamine

Endpoint summary

Description of key information

A NOAEL of 160 mg/kg bw/d for Caprinoguanamine was obtained for sub-acute toxicity from a Combined Repeated Dose Toxicity Study with the Reproduction / Developmental Toxicity Screening Test according to OECD guideline 422.

Key value for chemical safety assessment

Toxic effect type:

dose-dependent

Repeated dose toxicity: via oral route - systemic effects

Link to relevant study records

Reference

Reference 1

Endpoint:

short-term repeated dose toxicity: oral

Remarks:

Combined Repeated Dose Toxicity Study with the Reproductive/Developmental Toxicity Screening Study

Type of information:

experimental study

Adequacy of study:

key study

Study period:

2017-01-26 to 2017-09-20

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Reason / purpose for cross-reference:

reference to same study

Qualifier:

according to guideline

Guideline:

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

Version / remarks:

2016-06-29

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Limit test:

no

Species:

rat

Strain:

Sprague-Dawley

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Crl:CD(SD) rat /Charles River (UK) Ltd.
- Age at study initiation: Males: 69 to 76 days old, Females: 83 to 90 days old.
- Weight at study initiation: Males: 341 to 400 g, Females: 241 to 297 g.
- Housing: polycarbonate body with a stainless steel mesh lid. used during the acclimatization, pre-pairing, treatment, gestation, littering and lactation periods.
Grid bottomed polypropylene cages were used during pairing.
Number of animals per cage: Pre-pairing up to five animals of one sex; Pairing one male and one female; Males after mating up to five animals;
Gestation one female; Lactation one female + litter
- Diet (e.g. ad libitum): SDS VRF1 Certified pelleted ad libitum
- Water (e.g. ad libitum): ad libitum, from the public supply via polycarbonate bottles with sipper tubes.
- Acclimation period: Males: Six days before commencement of treatment ; Females: 20 days before commencement of treatment.

DETAILS OF FOOD AND WATER QUALITY:
The diet contained no added antibiotic or other chemotherapeutic or prophylactic agent.
Certificates of analysis for the diet are scrutinized and approved before any batch of diet was released for use.
Certificates of analysis were routinely provided by the water supplier.


ENVIRONMENTAL CONDITIONS
- Temperature (°C): 20-24ºC
- Humidity (%): 40-70%
- Air changes (per hr):
- Photoperiod (hrs dark / hrs light): 12 hours light : 12 hours dark

IN-LIFE DATES: From: 2017-05-03 To: 2017-07-02

Route of administration:

oral: gavage

Details on route of administration:

Oral gavage using a suitably graduated syringe and a rubber catheter inserted via the mouth.

Vehicle:

corn oil

Details on oral exposure:

- PREPARATION OF DOSING SOLUTIONS:

- VEHICLE
- Justification for use and choice of vehicle (if other than water): Standard vehicle according to guideline
- Concentration in vehicle: 8, 16, 32 mg/mL
- Amount of vehicle (if gavage): 5 mL

Analytical verification of doses or concentrations:

yes

Details on analytical verification of doses or concentrations:

The analytical procedure was successfully validated with respect to specificity of chromatographic analysis, limit of detection and quantification, linearity of detector response, repeatability, method accuracy and precision.
The homogeneity and stability was confirmed for Caprinoguanamine in corn oil formulations at nominal concentrations of 1 mg/mL and 200 mg/mL during distribution between the bottles, during magnetic stirring for 2 hours, ambient temperature storage for 1 day and refrigerated storage for up to 15 days.
The mean concentrations of Caprinoguanamine in test formulations analyzed for the study were within ±6% of nominal concentrations, confirming the accurate preparation of these formulations.

Duration of treatment / exposure:

Males: Two weeks before pairing up to necropsy after a minimum of five weeks of treatment (animals were killed in Week 6).
Females Two weeks before pairing, then throughout pairing and gestation until Day 13 of lactation.

Frequency of treatment:

Once daily at approximately the same time each day.
Animals were not dosed if parturition was in progress at the scheduled time of administration.

Dose / conc.:

0 mg/kg bw/day (nominal)

Dose / conc.:

40 mg/kg bw/day (nominal)

Dose / conc.:

80 mg/kg bw/day (nominal)

Dose / conc.:

160 mg/kg bw/day (nominal)

No. of animals per sex per dose:

10 males and 10 females

Control animals:

yes, concurrent vehicle

Details on study design:

- Dose selection rationale:
- Rationale for animal assignment (if not random):
Estrous cycles were evaluated prior to treatment. After 14 days evaluation, animals that failed to exhibit typical 4-5 day cycles were not allocated to the study.

On Day 1 of study all animals were weighed and body weights were reviewed by Study Management before dosing commenced to ensure that variations in
body weight of animals did not exceed ±20% of the mean for each sex. The groups were adjusted to reduce inter-/intra-group variation.

Positive control:

no

Observations and examinations performed and frequency:

CAGE SIDE OBSERVATIONS: Yes
- Time schedule: Animals were inspected visually at least twice daily for evidence of ill-health or reaction to treatment.
- Cage side observations: Any deviation from normal was recorded at the time in respect of nature and severity, date and time of onset, duration and progress of the observed condition, as appropriate.

DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: Before treatment commenced and during each week of treatment and on Days 0, 7, 14 and 20 after mating and Days 1, 6 and 12 of lactation,
detailed physical examination and arena observations were performed on each animal.

BODY WEIGHT: Yes
- Time schedule for examinations: males: Before dosing on the day treatment commenced (Day 1) and weekly thereafter. On the day prior to necropsy.
females: Before dosing on the day treatment commenced (Day 1) and weekly before pairing. Days 0, 7, 14 and 20 after mating. Day 1, 4, 7 and 13 of lactation. On the day prior to necropsy.

FOOD CONSUMPTION AND COMPOUND INTAKE: not applicable, no feeding study

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): not applicable, no drinking water study

OPHTHALMOSCOPIC EXAMINATION: No

HAEMATOLOGY: Yes
- Time schedule for collection of blood: at termination
- Anaesthetic used for blood collection: Yes, light general anesthesia induced by isoflurane
- Animals fasted: Yes; overnight
- How many animals: The five lowest numbered surviving males and the first five lactating females with a surviving litter, in each dose group.
- Parameters checked in table [No.1] were examined.

CLINICAL CHEMISTRY: Yes
- Time schedule for collection of blood: at termination
- Animals fasted: Yes overnight
- How many animals: The five lowest numbered surviving males and the first five lactating females with a surviving litter, in each dose group.
- Parameters checked in table [No.2] were examined.

URINALYSIS: No

NEUROBEHAVIOURAL EXAMINATION: Yes
- Time schedule for examinations: During Week 5 of treatment for males and at Day 7-9 of lactation for females
- Dose groups that were examined: five lowest numbered surviving males and the first five lactating females in each group
- Battery of functions tested: sensory activity / grip strength / motor activity

IMMUNOLOGY: No

THYROID HORMONE ANALYSIS.
- Time schedule for collection of blood: at termination, all surviving F0 adult males and females
- Anaesthetic used for blood collection: Yes, light general anesthesia induced by isoflurane
- Animals fasted: Yes; overnight
- How many animals: all surviving F0 adult males and females
Initially measurement of the serum samples of the adult males and the Day 13 offspring for thyroxine (T4) levels.
These examinations were undertaken and no effect of treatment was evident on the circulating levels of this hormone.
It was therefore not necessary to analyze any further samples for either T4 or TSH (thyroid stimulating hormone).

Sacrifice and pathology:

GROSS PATHOLOGY: Yes
All adult animals were subject to a detailed necropsy.
F0 males After Week 5 investigations completed.
F0 females failing to produce a viable litter: Day 25 after mating.
F0 females Day 14 of lactation.

Pathology procedures for the five lowest numbered surviving males and the first five lactating females with a surviving litter per group at scheduled termination [Table 3]

Pathology procedures for remaining F0 males and females per group [Table 4]

HISTOPATHOLOGY: Yes (Table 3 and 4)
Tissue samples were dehydrated, embedded in paraffin wax and sectioned at a nominal four to five micron thickness.
For bilateral organs, sections of both organs were prepared.
A single section was prepared from each of the remaining tissues required.

Full List: [Table 3] The five lowest numbered surviving males and the first five lactating females with a surviving litter in Groups 1 and 4 at scheduled termination.
Liver: The five lowest numbered surviving males in Groups 2 and 3.
Abnormalities only: All remaining F0 animals.

Statistics:

A parametric analysis was performed if Bartlett's test for variance homogeneity (Bartlett 1937) was not significant at the 1% level. For pretreatment data, analysis of variance was used to test for any group differences. Where this was significant (p<0.05) inter group comparisons using t-tests, with the error mean square from the one-way analysis of variance, were made. For all other comparisons the F1 approximate test was applied.
If the F1 approximate test for monotonicity of dose-response was not significant at the 1% level, Williams' test for a monotonic trend was applied.
If the F1 approximate test was significant, suggesting that the dose response was not monotone, Dunnett's test (Dunnett 1955, 1964) was performed instead. A non-parametric analysis was performed if Bartlett's test was still significant at the 1% level following both logarithmic and square-root transformations. For pretreatment data, Kruskal-Wallis’ test (Kruskal and Wallis 1952, 1953) was used to test for any group differences. Where this was significant (p<0.05) inter group comparisons using Wilcoxon rank sum tests (Wilcoxon 1945) were made. For all other comparisons the H1 approximate test, the non-parametric equivalent of the F1 test described above, was applied.
For grip strength, motor activity, clinical pathology data, if 75% of the data (across all groups) were the same value, for example c, Fisher’s exact tests (Fisher 1973) were performed.
For organ weight data, analysis of covariance was performed using terminal body weight as covariate (Angervall and Carlstrom, 1963), unless non-parametric methods were applied.
Significant differences between the groups compared were expressed at the 5% (p<0.05) or 1% (p<0.01) level.

Clinical signs:

no effects observed

Description (incidence and severity):

Parental (F0): Clinical condition, behaviour in the arena, sensory reactivity, grip strength and motor activity were unaffected by treatment.

Mortality:

no mortality observed

Description (incidence):

Parental (F0): No animals died prematurely

Body weight and weight changes:

no effects observed

Description (incidence and severity):

Parental (F0):
Male body weight gains during the last two weeks of treatment for animals receiving the test item at all doses were lower than controls.
This resulted in lower overall weight gains (Day 1 to 38) in these groups (96, 95 and 85% of control for males receiving 40, 80 or 160 mg/kg/day, respectively).
Females receiving 160 mg/kg/day had low body weight gains, when compared to controls, throughout the gestational period and subsequently a slightly lower gain throughout lactation.
These reductions in body weight gain were considered not of sufficient magnitude to be considered adverse.

Food efficiency:

not examined

Ophthalmological findings:

not examined

Haematological findings:

no effects observed

Description (incidence and severity):

Parental (F0):
Haematological investigations did not reveal any findings that were attributed to treatment

Clinical biochemistry findings:

no effects observed

Description (incidence and severity):

Parental (F0):
The biochemical examination of the blood plasma revealed slightly low plasma potassium, calcium, glucose, cholesterol and triglyceride concentrations in males receiving 40, 80 or 160 mg/kg/day.
Females receiving 40, 80 or 160 mg/kg/day had slightly higher plasma bile acid concentrations; these variations were in the absence of a dose dependent trend.

Urinalysis findings:

not examined

Behaviour (functional findings):

no effects observed

Description (incidence and severity):

Behaviour in the arena, sensory reactivity, grip strength and motor activity were unaffected by treatment.

Immunological findings:

not examined

Organ weight findings including organ / body weight ratios:

no effects observed

Description (incidence and severity):

When compared with the controls, the evaluation of organ weights of males killed after five weeks of treatment revealed the following non dose related changes which were considered of uncertain relationship to treatment.
The weight of the levator ani-bulbocavernosus muscle complex (LABC) was marginally but statistically significantly high in males given 160 mg/kg/day.
The weight of the thyroids/parathyroids was marginally but statistically significantly high in males given 80 or 160 mg/kg/day.

Gross pathological findings:

no effects observed

Description (incidence and severity):

The macroscopic examination performed after five weeks of treatment (males) or on Day 14 of lactation (females) did not reveal any test item-related changes.
The incidence and distribution of all findings were considered to be unrelated to treatment.

Histopathological findings: non-neoplastic:

effects observed, treatment-related

Description (incidence and severity):

Changes related to treatment with Caprinoguanamine were seen in the liver of males.
Minimal to moderate periportal hepatocellular vacuolation, was seen in most males at all doses of Caprinoguanamine, and at minimal to slight severity in two control males.

Histopathological findings: neoplastic:

no effects observed

Details on results:

The microscopic examination of the full list of examined tissues revealed that periportal hepatocellular vacuolation, typically a background change, was present at a higher incidence and severity in the livers of males receiving 40, 80 or 160 mg/kg/day of the test item. A mixed vacuolation was present in males given 160 mg/kg/day: in the periportal areas it was microvesicular, whereas at the edges of the liver lobes it was macrovesicular. In males given 40 or 80 mg/kg/day the vacuolation was more of the microvesicular type. Microvesicular lipidosis is usually indicative of more serious hepatic dysfunction but can also result from nutritional disturbances (Greaves 2007). There was no clear dose-dependency but a no observed effect level (NOEL) was not determined because the change was seen in males given the lowest dose of Caprinoguanamine. This change was considered not to represent an adverse effect of treatment since there was no effect on the clinical condition or survival of the animals and no signs of altered hepatic function as judged by clotting times and urea, protein and albumin levels.

Key result

Dose descriptor:

NOAEL

Effect level:

160 mg/kg bw/day (nominal)

Based on:

test mat.

Sex:

male/female

Basis for effect level:

other: highest tested dose

Key result

Critical effects observed:

no

Conclusions:

The no-observed-adverse-effect-level (NOAEL) for systemic toxicity was considered to be 160 mg/kg/day.

Executive summary:

In a Combined Repeated Dose Toxicity Study with the Reproduction / Developmental Toxicity Screening study according to OECD guideline 422, Caprinoguanamine (ca. 98 % a.i) was administered to 10 Sprague-Dawley rats /sex/ at dose levels of 0, 40, 80 and 160 mg/kg bw/day by gavage.

Males were treated daily for two weeks before pairing, up to necropsy after a minimum of five consecutive weeks. Females were treated daily for two weeks before pairing, throughout pairing, gestation and until Day 13 of lactation.

Parental (F0) responses

No animals died prematurely. Clinical condition, behaviour in the arena, sensory reactivity, grip strength and motor activity were unaffected by treatment. Salivation and chin rubbing were seen in association with dosing; such signs are common on studies dosed by the oral gavage route and are generally due to the taste of the formulations.

Male body weight gains during the last two weeks of treatment for animals receiving the test item at all doses were lower than controls. This resulted in lower overall weight gains (Day 1 to 38) in these groups (96, 95 and 85% of control for males receiving 40, 80 or 160 mg/kg/day, respectively). Females receiving 160 mg/kg/day had low body weight gains, when compared to controls, throughout the gestational period and subsequently a slightly lower gain throughout lactation. These reductions in body weight gain were considered not of sufficient magnitude to be considered adverse.

 

The food consumption of males receiving 160 mg/kg/day was slightly low in the first three weeks of treatment when compared with the controls. During the pre-pairing period, female food consumption was unaffected by treatment. However, during gestation, females receiving 160 mg/kg/day ate less food than the control group. Low food consumption was also seen in these females during lactation after Day 4. These minor variations in food consumption were considered non-adverse.

 

Estrous cyclicity, pre-coital interval, fertility, mating performance, gestation length and index were unaffected by the administration of Caprinoguanamine at all dose levels. There was no effect of treatment on the number of implantations or litter size.

Haematological investigations did not reveal any findings that were attributed to treatment.

The biochemical examination of the blood plasma revealed slightly low plasma potassium, calcium, glucose, cholesterol and triglyceride concentrations in males receiving 40, 80 or 160 mg/kg/day. Females receiving 40, 80 or 160 mg/kg/day had slightly higher plasma bile acid concentrations; these variations were in the absence of a dose dependent trend.

There was no effect of treatment on the circulating levels of thyroxine (T4) in adult males or in the Day 13 male and female offspring.

There were no changes in organ weights considered to be related to treatment

Macroscopic examination of the adult males and females did not reveal any test item-related changes, the incidence and distribution of all findings were considered to be unrelated to treatment.

Microscopic pathology revealed minimal to moderate periportal hepatocellular vacuolation in the liver of the majority of males receiving 40, 80 or 160 mg/kg/day. Similar findings were also recorded in two control males.

 

In conclusion, oral gavage administration of Caprinoguanamine for five weeks was generally well tolerated in males at dose levels of 40, 80 or 160 mg/kg/day but did result in a higher incidence and severity of periportal hepatocellular vacuolation in the liver at all dose levels when compared with the controls. Administration of Caprinoguanamine to females for two weeks before pairing, throughout gestation and up to Day 14 of lactation was also generally well tolerated with no test item related microscopic pathology changes detected.

 

In the context of this study, Caprinoguanamine showed no evidence of being an endocrine disruptor.

 

The no-observed-adverse-effect-level (NOAEL) for systemic toxicity was considered to be 160 mg/kg/day.

Reference 2

Endpoint:

short-term repeated dose toxicity: oral

Remarks:

Dose - Range - finder

Type of information:

experimental study

Adequacy of study:

supporting study

Study period:

2017-03-15 to 2017-04-11

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

study well documented, meets generally accepted scientific principles, acceptable for assessment

Qualifier:

no guideline required

Principles of method if other than guideline:

The purpose of this study was to assess the systemic toxic potential of Caprinoguanamine, a stabilizer, when administered to Sprague-Dawley (CD) rats by oral gavage for 14 days, and to aid in the selection of a high dose for a subsequent OECD 422 screening study.

GLP compliance:

no

Remarks:

Work performed generally followed Good Laboratory Practice principles

Limit test:

no

Species:

rat

Strain:

Sprague-Dawley

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source:
Crl:CD(SD) rat /Charles River (UK) Ltd.
- Age at study initiation:
Groups 1 to 4: males: 69 to 76 days, females: 83 to 90 days.
Group 5: males: 76 to 83 days, females: 90 to 97 days.
- Weight at study initiation:
Males: 340 to 426 g, Females: 217 to 317 g.
- Housing: polycarbonate body with a stainless steel mesh lid.
Number of animals per cage: Three of the same sex.
- Diet (e.g. ad libitum): SDS VRF1 Certified pelleted ad libitum
- Water (e.g. ad libitum): ad libitum, from the public supply via polycarbonate bottles with sipper tubes.
- Acclimation period: Six days before commencement of treatment ; group 5 13 days before commencement of treatment.

DETAILS OF FOOD AND WATER QUALITY:
Certificates of analysis for the diet were scrutinized and approved before any batch of diet was released for use.
Certificates of analysis are routinely provided by the water supplier.


ENVIRONMENTAL CONDITIONS
- Temperature (°C):
20-24ºC
- Humidity (%):
40-70%
- Air changes (per hr):

- Photoperiod (hrs dark / hrs light):
12 hours light : 12 hours dark

IN-LIFE DATES: From: 2017-03-15 To: 2017-04-11

Route of administration:

oral: gavage

Details on route of administration:

Oral gavage using a suitably graduated syringe and a rubber catheter inserted via the mouth.

Vehicle:

corn oil

Details on oral exposure:

- PREPARATION OF DOSING SOLUTIONS:

- VEHICLE
- Justification for use and choice of vehicle (if other than water): Standard vehicle according to guideline
- Concentration in vehicle: 25, 50, 100, 200 mg/mL
- Amount of vehicle (if gavage): 5 mL

Analytical verification of doses or concentrations:

no

Details on analytical verification of doses or concentrations:

No formulation analysis was performed on this study.

Duration of treatment / exposure:

14 days

Frequency of treatment:

daily

Dose / conc.:

0 mg/kg bw/day (nominal)

Dose / conc.:

125 mg/kg bw/day (nominal)

Dose / conc.:

250 mg/kg bw/day (nominal)

Dose / conc.:

500 mg/kg bw/day (nominal)

Dose / conc.:

1 000 mg/kg bw/day (nominal)

No. of animals per sex per dose:

three male and three female rats

Control animals:

yes, concurrent vehicle

Details on study design:

- Dose selection rationale:
The doses used in this study (0, 250, 500, 1000 and 125 mg/kg/day) were selected in conjunction with the Sponsor.

An acute oral toxicity study in the rat demonstrated that the acute median lethal oral dose
(LD50) of Caprinoguanamine was greater than 10000 mg/kg body weight.
Therefore the high dose for this study was 1000 mg/kg/day as this is the maximum dose level required for the subsequent OECD422 study.

Observations and examinations performed and frequency:

Animals were inspected visually at least twice daily for evidence of ill-health or reaction to treatment.
Cages were inspected daily for evidence of animal ill-health amongst the occupants.
Any deviation from normal was recorded at the time in respect of nature and severity, date and time of onset, duration and progress of the observed condition, as appropriate.

Clinical Signs
A detailed physical examination was performed, where appropriate, on Days 1, 4, 8, 11, 15, 18 and 22 on each animal, as appropriate, to monitor general health.

Mortality
A viability check was performed near the start and end of each working day. Animals were killed for reasons of animal welfare where necessary.

Body Weight
The weight of each animal was recorded three days before treatment commenced, on the day that treatment commenced, twice weekly throughout the study and before necropsy (Days -3, 1, 4, 8, 11, 15, 18 and 22 as appropriate).

Food Consumption
The weight of food supplied to each cage, that remaining and an estimate of any spilled was recorded for the three days before treatment commenced and twice weekly throughout the study
(Days -3 to -1, 1 to 3, 4 to 7, 8 to 10, 11 to 14, 15 to 17 and 18 to 21 as appropriate).

Water Consumption
Fluid intake was assessed by daily visual observation.

Sacrifice and pathology:

Necropsy
All animals were subject to a necropsy. The thoracic, abdominal and cranial cavities were opened. Any abnormality in the appearance or size of any organ and tissue (external and cut surface) was recorded and the required tissue samples preserved in fixative. The retained tissues were checked before disposal of the carcass.
Surviving animals were killed following 14 days of treatment (21 for Group 1).

The organs weighed and tissue samples fixed are detailed as follows:
Abnormalities, Kidneys, Lever, Spleen

Statistics:

No statistical analysis of the data was performed on this study.

Mortality:

mortality observed, treatment-related

It was concluded that the oral administration of Caprinoguanamine to Sprague Dawley rats at doses of 500 or 1000 mg/kg/day was not tolerated, with all animals at these dose levels being

killed for welfare reasons due to the severe signs seen after dosing on Day 1.

Although 250 mg/kg/day was generally tolerated, the adverse clinical signs seen at this dose level (piloerection, unsteadiness and limited use of the limbs) and the pallor of the liver seen

macroscopically were considered to preclude the use of this dose level for a study of longer duration.

At 125 mg/kg/day, low body weight gains were observed during the second week of treatment, with some degree of weight loss recorded for the majority of animals.

A high dose level of 160 mg/kg/day for the subsequent OECD 422 study with Caprinoguanamine is therefore considered likely to be tolerated, but will result in some toxicity.

Conclusions:

A high dose level of 160 mg/kg/day for the subsequent OECD422 study with Caprinoguanamine is therefore considered likely to be tolerated, but will result in some toxicity.

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed

Dose descriptor:

NOAEL

160 mg/kg bw/day

Study duration:

subacute

Species:

rat

Quality of whole database:

Data from a GLP compliant guideline studies with reliability 1.

Organ:

not specified

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

In a Combined Repeated Dose Toxicity Study with the Reproduction / Developmental Toxicity Screening study according to OECD guideline 422, Caprinoguanamine (ca. 98 % a.i) was administered to 10 Sprague-Dawley rats /sex/ at dose levels of 0, 40, 80 and 160 mg/kg bw/day by gavage.

Males were treated daily for two weeks before pairing, up to necropsy after a minimum of five consecutive weeks. Females were treated daily for two weeks before pairing, throughout pairing, gestation and until Day 13 of lactation.

Parental (F0) responses

No animals died prematurely. Clinical condition, behaviour in the arena, sensory reactivity, grip strength and motor activity were unaffected by treatment. Salivation and chin rubbing were seen in association with dosing; such signs are common on studies dosed by the oral gavage route and are generally due to the taste of the formulations.

Male body weight gains during the last two weeks of treatment for animals receiving the test item at all doses were lower than controls. This resulted in lower overall weight gains (Day 1 to 38) in these groups (96, 95 and 85% of control for males receiving 40, 80 or 160 mg/kg/day, respectively). Females receiving 160 mg/kg/day had low body weight gains, when compared to controls, throughout the gestational period and subsequently a slightly lower gain throughout lactation. These reductions in body weight gain were considered not of sufficient magnitude to be considered adverse.

 

The food consumption of males receiving 160 mg/kg/day was slightly low in the first three weeks of treatment when compared with the controls. During the pre-pairing period, female food consumption was unaffected by treatment. However, during gestation, females receiving 160 mg/kg/day ate less food than the control group. Low food consumption was also seen in these females during lactation after Day 4. These minor variations in food consumption were considered non-adverse.

 

Estrous cyclicity, pre-coital interval, fertility, mating performance, gestation length and index were unaffected by the administration of Caprinoguanamine at all dose levels. There was no effect of treatment on the number of implantations or litter size.

Haematological investigations did not reveal any findings that were attributed to treatment.

The biochemical examination of the blood plasma revealed slightly low plasma potassium, calcium, glucose, cholesterol and triglyceride concentrations in males receiving 40, 80 or 160 mg/kg/day. Females receiving 40, 80 or 160 mg/kg/day had slightly higher plasma bile acid concentrations; these variations were in the absence of a dose dependent trend.

There was no effect of treatment on the circulating levels of thyroxine (T4) in adult males or in the Day 13 male and female offspring.

There were no changes in organ weights considered to be related to treatment

Macroscopic examination of the adult males and females did not reveal any test item-related changes, the incidence and distribution of all findings were considered to be unrelated to treatment.

Microscopic pathology revealed minimal to moderate periportal hepatocellular vacuolation in the liver of the majority of males receiving 40, 80 or 160 mg/kg/day. Similar findings were also recorded in two control males.

 

In conclusion, oral gavage administration of Caprinoguanamine for five weeks was generally well tolerated in males at dose levels of 40, 80 or 160 mg/kg/day but did result in a higher incidence and severity of periportal hepatocellular vacuolation in the liver at all dose levels when compared with the controls. Administration of Caprinoguanamine to females for two weeks before pairing, throughout gestation and up to Day 14 of lactation was also generally well tolerated with no test item related microscopic pathology changes detected.

 

In the context of this study, Caprinoguanamine showed no evidence of being an endocrine disruptor.

 

The no-observed-adverse-effect-level (NOAEL) for systemic toxicity was considered to be 160 mg/kg/day.

Justification for classification or non-classification

Repeated dose toxicity of Caprinoguanamine was assessed based on data from a Combined Repeated Dose Toxicity Study with the Reproduction / Developmental Toxicity Screening study.

The no-observed-adverse-effect-level (NOAEL) for systemic toxicity was considered to be 160 mg/kg/day, the highest tested dose.

Evaluating information from the sub-acute toxicity study a classification with STOT-RE according to GHS Regulation EC No 1272/2008 for Caprinoguanamine is considered not warrant.