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Toxicity to reproduction

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Administrative data

Endpoint:
two-generation reproductive toxicity
Remarks:
based on test type (migrated information)
Type of information:
migrated information: read-across from supporting substance (structural analogue or surrogate)
Adequacy of study:
key study
Study period:
2007-Jan-09 trough 2007-May-22
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: GLP guideline study

Data source

Referenceopen allclose all

Reference Type:
study report
Title:
Unnamed
Year:
2008
Report Date:
2008
Reference Type:
study report
Title:
Unnamed
Year:
2007
Report Date:
2007

Materials and methods

Test guidelineopen allclose all
Qualifier:
according to
Guideline:
OECD Guideline 416 (Two-Generation Reproduction Toxicity Study)
Deviations:
no
Qualifier:
according to
Guideline:
EPA OPPTS 870.3800 (Reproduction and Fertility Effects)
Deviations:
no
Qualifier:
according to
Guideline:
EU Method B.35 (Two-Generation Reproduction Toxicity Test)
Deviations:
no
GLP compliance:
yes (incl. certificate)
Limit test:
no

Test material

Reference
Name:
Unnamed
Type:
Constituent
Details on test material:
- Name of test material (as cited in study report): Sodium formate
- Physical state: solid
- Analytical purity: 100%

Test animals

Species:
rat
Strain:
Wistar
Sex:
male/female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Charles River Laboratories, Research Models and Services, Germany GmbH.
- Age at study initiation: (P) 7 wks
- Weight at study initiation: (P) Males: men 108.0 g; Females: mean 96.0 g
- Fasting period before study: ovenight; feed study
- Housing: individually in Macrolon type M III cages supplied by Becker & Co., Castrop-Rauxel, Germany (floor area of about 800 cm²)
- Diet: ground Kliba maintenance diet mouse/rat “GLP” meal, supplied by Provimi Kliba SA, Kaiseraugst, Switzerland, was available ad libitum
throughout the study.
- Water: tap water ad libitum
- Acclimation period: 7 days


ENVIRONMENTAL CONDITIONS
- Temperature (°C): 20-24
- Humidity (%): 30-70
- Air changes (per hr): 10-15
- Photoperiod (hrs dark / hrs light): 12/12

Administration / exposure

Route of administration:
oral: feed
Vehicle:
other: food
Details on exposure:

DIET PREPARATION
- Rate of preparation of diet (frequency): weekly
- Mixing appropriate amounts with (Type of food): ground standard diet
- Storage temperature: room temperature
Details on mating procedure:
- M/F ratio per cage: 1:1
- Length of cohabitation: maximum two weeks
- Proof of pregnancy: sperm in vaginal smear referred to as day 0 of pregnancy
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
Commercial test kit "Ameisensäure"; enzymatic test with UV/vis detection at 340 nm. Details: Study Part III, p. 882
Duration of treatment / exposure:
Exposure period: throughout the study
F0 and F1 generation: 75 days premating exposure; during mating and gestation until post natal day (PND) 21
Frequency of treatment:
daily
Details on study schedule:
- F1 parental animals not mated until 10 weeks after selected from the F1 litters.
Doses / concentrations
Remarks:
Doses / Concentrations:
0, 100, 300 and 1000 mg/kg body weight/day
Basis:
nominal in diet
No. of animals per sex per dose:
25
Control animals:
yes, concurrent vehicle
Details on study design:
- Dose selection rationale: based on results of preceding studies

Examinations

Parental animals: Observations and examinations:
CAGE SIDE OBSERVATIONS: Yes
- Time schedule: A check for moribund or dead animals was made twice daily on working days or once daily
(Saturday, Sunday or on public holidays).


DETAILED CLINICAL OBSERVATIONS: Yes
- Tim eschedule: daily


BODY WEIGHT: Yes
- Time schedule for examinations: weekly


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: Yes
Estrous cyclicity (parental animals):
Estrous cycle length and normality were evaluated daily for all F0 and F1 female parental rats for a minimum of 3 weeks prior to mating. The evaluations were continued throughout the mating period until the female exhibited evidence of mating. Moreover, at the scheduled necropsy a vaginal smear was microscopically examined to determine the stage of the estrous cycle for each F0 and F1 female.
Sperm parameters (parental animals):
Parameters examined in [all/P/F1/F2] male parental generations:
testis weight, epididymis weight, sperm count in testes and in epididymides, sperm motility, sperm morphology
Litter observations:
STANDARDISATION OF LITTERS
- Performed on day 4 postpartum: yes
- If yes, maximum of 8 pups/litter (4/sex/litter as nearly as possible); excess pups were killed and discarded.


PARAMETERS EXAMINED
The following parameters were examined in F1 and F2 offspring:
number and sex of pups, stillbirths, live births, postnatal mortality, presence of gross anomalies, weight at birth and weight gain, physical or behavioural abnormalities, sexual maturation (vaginal openng, preputial separation).


GROSS EXAMINATION OF DEAD PUPS:
yes, for external and internal abnormalities; possible cause of death was not determined for pups born or found dead. Determinationof pup organweight (brain, spleen, thymus).
Postmortem examinations (parental animals):
SACRIFICE
F0 and F1 animals:
- Male animals: All surviving animals after the F1 or F2 pups were weaned (PND 21).
- Maternal animals: All surviving animals after the F1 or F2 pups were weaned (PND 21).


GROSS NECROPSY
- Gross necropsy consisted of external and internal examinations including the cervical, thoracic, and abdominal viscera.

HISTOPATHOLOGY / ORGAN WEIGHTS
The tissues indicated in section 3.9.2 (page 51) were weighed and those in section 3.9.3 (page 52) were prepared for microscopic examination.
Tissues examined by light microscopy are tabulated in section 3.9.4.
Postmortem examinations (offspring):
SACRIFICE
- The F1 offspring not selected as parental animals were sacrificed at 4 days of age and all F2 offspring were sacrificed at 21 days of age.
- These animals were subjected to postmortem examinations (macroscopic and/or microscopic examination) as follows:


GROSS NECROPSY
- Gross necropsy consisted of external and internal examinations including the cervical, thoracic, and abdominal viscera.


HISTOPATHOLOGY / ORGAN WEIGTHS
Brain, spleen, and thymus were weighed.
Statistics:
Extensive statistical evaaluations included Dunnett test; Fisher's exact test; Wilcoxon test; Kruskal-Wallis test (cf. report, section 3.8.3)
Reproductive indices:
Male mating index; male fertility index. Female mating index; female fertility index; gestation index. Live birth index; postimplantation loss.
Offspring viability indices:
Viability index day 4. Lactation index (viability on day 21)

Results and discussion

Results: P0 (first parental animals)

General toxicity (P0)

Clinical signs:
no effects observed
Body weight and weight changes:
no effects observed
Food consumption and compound intake (if feeding study):
no effects observed
Organ weight findings including organ / body weight ratios:
no effects observed
Gross pathological findings:
no effects observed
Histopathological findings: non-neoplastic:
no effects observed
Other effects:
effects observed, treatment-related

Reproductive function / performance (P0)

Reproductive function: estrous cycle:
no effects observed
Reproductive function: sperm measures:
no effects observed
Reproductive performance:
no effects observed

Details on results (P0)

CLINICAL SIGNS AND MORTALITY (PARENTAL ANIMALS)
No findings in F0 and and F1 parental animals noted.


BODY WEIGHT AND FOOD CONSUMPTION (PARENTAL ANIMALS)
No changes in F0 and and F1 parental animals noted.

TEST SUBSTANCE INTAKE (PARENTAL ANIMALS)
The mean test substance intake was close to the nominal values:
F0 males: 93 / 282 / 939 mg/kg bw/day in treated groups
F0 females: 97 / 291 / 977 mg/kg bw/day in treated groups

F1 males: 91 / 266 / 949 mg/kg bw/day in treated groups
F2 females: 98 / 295 / 980 mg/kg bw/day in treated groups


REPRODUCTIVE FUNCTION: ESTROUS CYCLE (PARENTAL ANIMALS)
No effect in F0 females (section 4.2.1.6) or F1 females (section 4.2.36)


REPRODUCTIVE FUNCTION: SPERM MEASURES (PARENTAL ANIMALS)
No treatment-related effct in F0 males (section 4.2.1.8) or F1 males (section 4.2.3.8)

REPRODUCTIVE PERFORMANCE (PARENTAL ANIMALS)
No effect in male or female F0 and F1 animals

F0 animals:
Males:
The male mating index varied between 96% in the control and 100% in the test groups.
The male fertility index ranged between 92% and 100% without showing any effect of dosing.
Females:
The female mating index calculated after the mating period for F1 litter was 96% in the control group and 100% in test groups 01-03 (100, 300 and 1000 mg/kg bw/d).
The mean duration until sperm was detected (GD 0) varied between 4.4 and 5.6 days without any relation to dosing. The fertility index was 96% in test group 01, 92% in test group 02 and 100% in test groups 00 and 03. The mean duration of gestation varied between 22.0 and 22.2 days without any relation to dosing.
The gestation index was 100% for all test groups, indicating that all pregnant F0 females had live F1 pups in their litters. There were no statistically significant differences in postimplantation loss between the test groups and the control (see Tab. IA-028). The live birth index was 99%, 99%, 100% and 99% in test groups 00-03.

F1 parental aimals
Males:
The male mating index was 96% in the low-dose group and 100% in the remaining test groups.
The male fertility index ranged between 92% and 100% without showing any effect of dosing.

Females: The female mating index for the F2 litter was 96% in test group 11 (100 mg/kg bw/d) and 100% in test groups 10, 12 and 13 (0, 300 and 1000 mg/kg bw/d).
The fertility index was 92% in the control and 100% in the remaining test groups.
The gestation index was 100% in all test groups, indicating that all pregnant F1 females had live F2 pups in their litters. The mean duration of gestation was very similar in all F1 test groups and varied between 21.8 and 22.0 days, with no relation to dosing.
There were no statistically significant differences of postimplantation loss between the test and the control groups.
The live birth index varied between 99% (test group 10) and 100% (test groups 11, 12 and 13).
ORGAN WEIGHTS (PARENTAL ANIMALS)
F0 animals:
Mean absolute organ weight parameters did not show significant differences compared to the control groups. In females, the mean relative weights of the brain, kidneys, and ovaries significantly changed (up to p<0.01), but were attributed to the slight reduction of the terminal body weight.

F1 animals:
Some significant changes (increase and reduction) of the absolute and relative weight were seen (kidneys, liver, pituitary gland, cauda epididymis affected; cf. section 4.3.2). There was no relation to dose or histopathological correlate.

GROSS PATHOLOGY and HISTOPATHOLOGY (PARENTAL ANIMALS)
F0 animals:
No changes noted (section 4.3.1)
F1 animals:
No changes apart from incidental glandular stomach erosion/ulcer in one control and eight mid dose animals.


OTHER FINDINGS (PARENTAL ANIMALS)

Sperm parameters: unchanged.
Estrous cycle data: no treatment-related change noted.

Effect levels (P0)

Dose descriptor:
NOAEL
Remarks:
reproduction
Effect level:
1 000 mg/kg bw/day
Sex:
male/female
Basis for effect level:
other: No adverse effect on reproductive performance and reproductive organs of F0 animals.

Results: F1 generation

General toxicity (F1)

Clinical signs:
no effects observed
Mortality / viability:
no mortality observed
Body weight and weight changes:
no effects observed
Sexual maturation:
no effects observed
Organ weight findings including organ / body weight ratios:
no effects observed
Gross pathological findings:
no effects observed
Histopathological findings:
no effects observed

Details on results (F1)

VIABILITY (OFFSPRING)

F1 pups:
The viability index indicating pup mortality during early lactation (PND 0-4) varied between 98% (test group 03 – 1000 mg/kg bw/d) and 99% (test groups 00, 01 and 02 – 0, 100 and 300 mg/kg bw/d).
The lactation index indicating pup mortality on PND 4-21 varied between 99% (test group 02 – 300 mg/kg bw/d) and 100% (test groups 00, 01 and 03 – 0, 100 and 1000 mg/kg bw/d).

F2 pups:
The viability index indicating pup mortality during early lactation (PND 0-4) varied between 99% (test groups 11, 12 and 13 – 100, 300 and 1000 mg/kg bw/d) and 100% (control group).
The lactation index indicating pup mortality on PND 4-21 varied between 98% (control group) and 100% (test groups 11, 12 and 13 – 100, 300 and 1000 mg/kg bw/d).

CLINICAL SIGNS (OFFSPRING)
There were no treatment related clinical signs in F0 and F1 pups.

BODY WEIGHT (OFFSPRING)
Mean body weights and mean body weight gain of the male and female F1 (Figure page 69) and F2 pups (Figure page 85) were not influenced by the test substance during the entire lactation period. All values in test groups at 100, 300 and 1000 mg/kg bw/d were comparable with the concurrent control group.

SEXUAL MATURATION (OFFSPRING)
F1 pups:
Males, preputial separation: the mean number of days to reach the separation was 41.4, 41.5, 41.2, and 42.0 days in controls and 100, 300 and 1000 mg/kg body weight/day test groups, indicating that the test substance did not influence male sexual maturation.
Females, vaginal opening: the mean number of days to reach the criterion amounted to 31.3, 30.8, 31.4, and 31.9 days in controls and 100, 300 and 1000 mg/kg body weight/day test groups, respectively, indicating that female sexual maturation was not influenced by the test substance.

F2 pups: not determined

ORGAN WEIGHTS (OFFSPRING)
F1 pups: mean absolute and relative pup organ weights did not show statistically significant differences to the control group, withth exception of statistically significantly increased absolute brain weights in groups at 100 and 300 mg/kg bw/day. Relative brain weight was not affected, and there was no dose relationship. Therefore, this finding was not considered to be of toxicological relevance.
F2 pups: mean absolute and relative pup organ weights did not show statistically significant differences to the control group, withth exception of statistically significantly decreased absolute and relative spleen weight in high dose males. Females were not affected. This finding was not considered to be of toxicological relevance.


GROSS PATHOLOGY (OFFSPRING)
There were spontaneous findings at necropsy of the F1 pups (Part III, tables IA-039 through IA-042) and F2 pups (Part III, tables IA-084 through IA-086), but these were without relation to dosing and/or could be found in the historical control data at comparable or even higher incidences.


OTHER FINDINGS (OFFSPRING)
Sex ratio was unchanged in the treated F1 and F2 pups.

Effect levels (F1)

Dose descriptor:
NOAEL
Remarks:
development
Generation:
F1
Effect level:
1 000 mg/kg bw/day
Sex:
male/female
Basis for effect level:
other: No effects on pre- and postnatal survival, body weights, no clinical or necropsy findings.

Overall reproductive toxicity

Reproductive effects observed:
not specified

Applicant's summary and conclusion

Conclusions:
Sodium formate was not a reproduction toxicant in a 2-generation rat feeding study.
Executive summary:

The fertility toxicity potential of sodium formate was examined in a 2-Generation feed study using male and female Wistar rats (25/sex and dose) at dose levels of 0, 100, 300, and 1000 mg/kg bw/day. The study was conducted in accordance with current test guidelines including OECD guideline No. 416, and under GLP conditions.

 

Test substance consumption

The mean test substance intake was close to the nominal values:

Nominal dose
(mg/kg bw/day)

100

300

1000

F0 males

93.2

282

939.1

F0 females
- premating
- gestation


96.8
105.4


290.9
323.9


976.9
1081.4

F1 males

90.7

265.6

948.7

F1 females
- premating
- gestation


97.9
108.1


294.8
276.6


980.1
931.3

                                              

 

Clinical and pathological findings in parental animals

There were no clinical signs of toxicity or mortalities in any of the F0 or F1 parental dose groups. Food consumption and body weights were comparable to that of the concurrent controls. Necropsy and pathology revealed no gross findings or organ weight changes that could be treatment related.

 

Reproduction parameters in F0 and F1 parental animals
There were no indications that sodium formate adversely affected fertility or reproductive performance of the F0 and F1 parental animals at dose levels as high as 1000 mg/kg body weight/day. Mating behavior, conception, gestation, parturition, lactation and weaning as well as sexual organ weights and gross findings of these organs were comparable between the rats of the test substance-treated test groups and the corresponding controls, and ranged within the historical control data of the test facility. There were no effects on male and female reproduction organs. Sperm parameters and estrous cycle were not affected.

Reproduction and developmental parameters in F1 pups

No test substance induced signs of developmental toxicity were noted in the progeny of the F0 and F1 parents at dose levels as high as 1000 mg/kg body weight/day. The number of delivered pups/litter, the sex ratio, their postnatal survival on days 4 and 21 after parturition, their body weights, and their sexual maturation remained unaffected by the test substance.

 

Clinical and pathological findings in F1 and F2 pups

-      Clinical and/or gross necropsy examinations of the F1 and F2 pups revealed only findings which were considered to be spontaneous in nature.The type and incidence offindings was within the range of the concurrent and/or the historical controls. (BASF SE, 2008).

 

Based on the above, the NOAEL values were as follows:

-      NOAEL 1000 mg/kg bw/day for general systemic toxicity for F0 and F1 parental animals

-      NOAEL 1000 mg/kg bw/day for fertility and reproductive performance for the F0 and F1 parental rats

-      NOAEL 1000 mg/kg bw/day for developmental toxicity, in the F1 and F2 progeny.

 

 

The study is considered to be valid and can be used for assessment of this endpoint.

 

For read across purposes, the NOAEL for the formate anion may be calculated, taking into account formula weights. The calculation (1000 mg sodium formate/kg /69 x 45 = 650 mg/kg bw/day) gives a NOAEL of approx. 650 mg formate/kg bw/day.