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Description of key information

Key value for chemical safety assessment

Repeated dose toxicity: via oral route - systemic effects

Link to relevant study records
Reference
Endpoint:
sub-chronic toxicity: oral
Type of information:
experimental study
Adequacy of study:
key study
Study period:
January 1987 - February 1988
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: Comparable to guideline study
Reason / purpose:
reference to same study
Qualifier:
no guideline followed
Principles of method if other than guideline:
30 male and 30 female Sprague-Dawley rats per group and generation (P and F1) were treated with zinc naphthenate. The dosage groups were as follows: 0, 500, 1000 and 5000 ppm in the diet. The vehicle was corn oil. In addition, a vehicle control group was also tested. The animals were treated 10 weeks prior to mating (males/females), during mating (males/females), gestation (females) and lactation (females).
GLP compliance:
yes
Remarks:
US-GLP (Title 40, Code of Federal Regulations, 1985 rev., Part 160, Good Laboratory Practice Standards
Limit test:
no
Species:
rat
Strain:
Sprague-Dawley
Sex:
male/female
Details on test animals and environmental conditions:
TEST ANIMALS - Sprague Dawley COBS, CD rats
- Source: Charles River Laboratories of Wilmington, Massachusetts.
- Age at study initiation: 7 weeks of age
- Housing: animals were kept three per cage,
- Diet (ad libitum): certified rodent ration (Zeigler Bros., Inc., Gardners, Pennsylvania). The ration arrived in the form of lab block and was ground to a uniform consistency.
- Water (ad libitum): water
- Quarantine period: a 2-week quarantine; Body weights and feed consumption were monitored during this pretreatment period.

ENVIRONMENTAL CONDITIONS
- Temperature: 21.1°C (18.3 - 23.9°C)
- Relative humidity: 50% (40% - 60%)
- Photoperiod (hrs dark / hrs light): 12/12
Route of administration:
oral: feed
Vehicle:
corn oil
Remarks:
Mazola
Details on oral exposure:
DIET PREPARATION
Feed was prepared on a weekly basis by dissolving the appropriate amount of zinc naphthenate in corn oil with the aid of heat, while keeping the total volume of corn oil/zinc naphthenate for each dosage group constant. These solutions were, in turn, poured into ground rodent ration and thoroughly blended with a mechanical mixer.
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
To ensure homogeneity and accuracy of the preparation, samples were extracted from each batch of feed and analysed for zinc content by atomic absorption.
Compound concentration in feed remained at a constant level throughout the entire study with no attempts to adjust for body weight gains.
Duration of treatment / exposure:
1) P generation:
Males:
- 10 weeks prior to mating
- throughout mating until necropsy
Females:
- 10 weeks prior to mating
- throughout mating, gestation and lactation

2) F1 generation:
Males:
- 10 weeks prior to mating starting after weaning
- throughout mating until necropsy
Females:
- 10 weeks prior to mating
- throughout mating, gestation and lactation
Frequency of treatment:
ad libitum
Remarks:
Doses / Concentrations:
500ppm (= 25mg/kg bw/day)
Basis:
nominal in diet
Remarks:
Doses / Concentrations:
1000ppm(= 50mg/kg bw/day)
Basis:
nominal in diet
Remarks:
Doses / Concentrations:
5000ppm(= 250mg/kg bw/day)
Basis:
nominal in diet
No. of animals per sex per dose:
P generation: 30 males / 30 females
F1 generation: 30 males / 30 females
Control animals:
yes, concurrent vehicle
Details on study design:
PILOT STUDY
The dosages of zinc naphthenate in the main study were based on the previously conducted pilot study which indicated effects between 2,700 ppm and 5,000 ppm in both parental sexes and their offspring.
The pilot study is described as follows:
A pilot study was conducted using 72 Sprague Dawley COBS, CD rats (six groups, each of six male and six female). Dosages for the study were calculated on a percent diet basis and were derived from expected food consumption and toxic signs seen in the acute oral studies. The concentration of zinc naphthenate in the feed ranged from 0.13 percent (1,300 ppm) in the low dosage group to 2.10 percent (21,000 ppm) in the high dosage group.

Results:
Male and female rats receiving 1 and 2 percent zinc naphthenate in their diet (10,000 ppm and 20,000 ppm) showed reduced body weights following 4 weeks of compound administration. Both sexes of rats in these groups also became lethargic and experienced urogenital staining from compound
consumption. Mating of rats within their respective groups indicated the 2 percent zinc naphthenate group required twice the mating time as did the other five dosage groups. Litters born of the highest two dosage groups (one and two percent) experienced reduced birth numbers and reduced total birth weights. At the time of weaning, offspring numbers and weights for these high dosage group litters were greatly reduced when compared to
litters of lower dosage groups and controls.
Positive control:
not stated
Observations and examinations performed and frequency:
1) P GENERATION

CAGE SIDE OBSERVATIONS: Yes
- Time schedule: animals were checked daily for toxic signs during the 10 weeks before mating.

DETAILED CLINICAL OBSERVATIONS: No

BODY WEIGHT: Yes
- Time schedule for examinations: body weights were recorded three times per week during the 10 weeks before mating.

FOOD CONSUMPTION AND COMPOUND INTAKE (if feeding study):
- Food consumption for each animal determined and mean daily diet consumption calculated as g food/kg body weight/day: Yes
- Compound intake calculated as time-weighted averages from the consumption and body weight gain data: No

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: No
HAEMATOLOGY: No
CLINICAL CHEMISTRY: No
URINALYSIS: No
NEUROBEHAVIOURAL EXAMINATION: No


2) F1 GENERATION

CAGE SIDE OBSERVATIONS: Yes
- Time schedule: daily during the 10 week exposure period prior to mating.

DETAILED CLINICAL OBSERVATIONS: No

BODY WEIGHT: Yes
- Time schedule for examinations: three times a week during the 10 week exposure period prior to mating

FOOD CONSUMPTION AND COMPOUND INTAKE (if feeding study):
- Food consumption for each animal determined and mean daily diet consumption calculated as g food/kg body weight/day: Yes
- Compound intake calculated as time-weighted averages from the consumption and body weight gain data: No data

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: No
HAEMATOLOGY: No
CLINICAL CHEMISTRY: No
URINALYSIS: No
NEUROBEHAVIOURAL EXAMINATION: No
Sacrifice and pathology:
1) P GENERATION

GROSS PATHOLOGY: Yes
HISTOPATHOLOGY: Yes
- Male animals: animals were euthanized by CO2, examined grossly, and tissues removed (testes, epididymides, seminal vesicles, prostate, pituitary, liver, kidneys), for histopathologic examination
- Maternal animals: the dams were sacrificed by CO2 at the time of weaning of the F1 generation. All dams were examined grossly while the vagina, uterus, ovaries, pituitary gland, liver, and kidneys were removed for histopathologic examination.

2) F1 GENERATION
- Male animals: following the 3-week mating period, F1 males were necropsied, taking sex and target organs for histopathologic examination.
- Maternal animals: F1 dams had organs taken for histopathologic examination.

In addition to the periodic sacrifices mentioned above, any animal dying spontaneously or for some reason removed from the study at an early date was submitted for gross necropsy. Gross necropsies consisted of the examination of all external surfaces, orifices, brain and spinal cord, thoracic, abdominal and pelvic cavities, and organs therein.
Clinical signs:
effects observed, treatment-related
Mortality:
mortality observed, treatment-related
Body weight and weight changes:
effects observed, treatment-related
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:
not examined
Clinical biochemistry findings:
not examined
Urinalysis findings:
not examined
Behaviour (functional findings):
not examined
Organ weight findings including organ / body weight ratios:
not examined
Gross pathological findings:
not specified
Histopathological findings: non-neoplastic:
effects observed, treatment-related
Histopathological findings: neoplastic:
no effects observed
Details on results:
1) P GENERATION

CLINICAL SIGNS AND MORTALITY
Male rats in the 0.5 percent dosage group appeared to become lethargic after week 9 of the dosing period. Alopecia was prevalent in the mid- and high-dosage animals of both sexes.
One male rat from the 0.1 percent dosage group was found cannibalised while the rats were group housed, prior to mating. Fighting was the apparent cause of death.

BODY WEIGHT AND WEIGHT GAIN
The P generation female rats in the 0.5 percent group (5000 ppm) experienced significantly reduced body weights by week 3 of compound administration, and continued through week 10. The control, low, and mid-dosage groups gained at similar rates.
Male rats fed a diet of 5000 ppm zinc naphthenate demonstrated significantly decreased body weights by week 8 of the 10 week dosing period.

FOOD CONSUMPTION AND COMPOUND INTAKE (if feeding study)
Weekly monitoring of feed consumption during the 10 week exposure period indicated all female rats ate similar portions, regardless of the dosage group. The same was also true for male rats, regardless of the zinc naphthenate concentration in the feed.

HISTOPATHOLOGY: NON-NEOPLASTIC
Histopathological examination of P generation rat tissues revealed compound-related lesions in the kidneys of male rats of the 0.5 percent dosage group. These lesions consisted of accumulations of amorphous to slightly granular, lightly eosinophilic material in the lumina of renal tubules, particularly near the corticomedullary junction. The term "nephrosis" was used to describe the lesions which were considered microscopically distinct from the intraluminal accumulations of homogenous proteinic material associated with the spontaneous degenerative nephropathy syndrome of rats. "Epithelial regeneration" of the renal tubules was evident, which may be assoicated with a variety of degenerative processes, especially the spontaneous degenerative nephropathy syndrome, mentioned above. However the increased incidence of this regeneration observed in the high-dosage rats suggests a compound-related effect. These lesions were not so advanced as to expect a clinically detectable effect on renal function. Lower concentrations of compound did not produce these effects, and all other findings were considered incidental.

2) F1 GENERATION

CLINICAL SIGNS AND MORTALITY
One dam, a control group animal, was found dead at approximately 17 days of pregnancy, and was submitted for necropsy. No apparent reason could be determined for the cause of death.

BODY WEIGHT AND WEIGHT GAIN
Body weights of female rats in the 0.5 percent dosage group were significantly depressed from the first week of the dosing period through week 10, when compared to the control group weights. Elevated body weights were observed in the 0.05 percent group from weeks 5 through 10.
As with the female rats, male rats in the 0.5 percent dosage group experienced significantly reduced lowered body weights throughout the pre-mating period. Again, male rats in the 0.05 percent group exceeded the body weights of the control and mid-dosage groups during weeks 7 through 10.

FOOD CONSUMPTION
Feed consumption figures during the dosing period revealed that all groups of rats, both male and female, ate similar portions of chow, and were not influenced by compound concentrations.

HISTOPATHOLOGY: NON-NEOPLASTIC
Histopathological examination of tissues form the F1 generation rats again showed "nephrosis " and " tubular regeneration" in the kidneys of a small number of male rats which was apparently associated with compound administration. All other findings were considered incidental or associated with spontaneous disease complexes of rats.
Key result
Dose descriptor:
NOAEL
Remarks:
(P Generation)
Effect level:
50 mg/kg bw/day (actual dose received)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
other: NOAEL for general toxicity based on effects on body weights.
Key result
Dose descriptor:
NOAEL
Remarks:
(F1 Generation)
Effect level:
50 mg/kg bw/day (actual dose received)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
other: NOAEL for general toxicity based on effects on body weights.
Key result
Critical effects observed:
not specified
Conclusions:
NOAEL general toxicity: 1000 ppm (corresponding to 50 mg/kw bw/day) based on effects on body weight.

P generation male and female rats fed with a high dose diet containing 5000 ppm zinc naphthenate showed body weight depression even though food consumption was comparable to other groups.
Lethargy was observed in males of the 5000 ppm group.

Endpoint conclusion
Endpoint conclusion:
adverse effect observed
Dose descriptor:
NOAEL
50 mg/kg bw/day
Study duration:
chronic
Species:
rat

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

One 2 -generation reproductive toxicity study in rats is available for zinc naphthenate. The study is considered relevant and reliable without restrictions.

P generation male and female rats fed with a high dose diet containing 5000 ppm zinc naphthenate showed slight body weight depression even though food consumption was comparable to other groups. Lethargy was observed in males of the 5000 ppm group. P generation male and female rats fed with a high dose diet containing 5000 ppm zinc naphthenate showed body weight depression even though food consumption was comparable to other groups. Although no full histopathological examination was performed on all organs of the test animals, the effects on body weight observed in the high dose group animals are considered as mild signs of general systemic toxicity, suitable for setting a no observed adverse effect level. The NOAEL for systemic toxicity is 1000 ppm (corresponding to 50 mg/kw bw/day) based on effects on body weight.

In conclusion, the NOAEL for systemic toxicity in this 2 -generation reproductive toxicity study also covers the endpoint for repeated dose toxicity, thus can be used to fulfil the data requirements as laid down in regulation (EC) 1907/2006. The NOAEL is used as point of departure in the DNEL derivation for naphthenic acid, zinc salts.

Justification for classification or non-classification