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Toxicity to reproduction

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Administrative data

Endpoint:
two-generation reproductive toxicity
Type of information:
experimental study
Adequacy of study:
key study
Study period:
2009
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
comparable to guideline study

Data source

Reference
Reference Type:
publication
Title:
Unnamed
Year:
2009

Materials and methods

Test guidelineopen allclose all
Qualifier:
equivalent or similar to guideline
Guideline:
OECD Guideline 416 (Two-Generation Reproduction Toxicity Study)
Deviations:
no
Qualifier:
according to guideline
Guideline:
other: protocol approved by the Institutional Animal Ethics Committee (Protocol No. P/4489/RT-DT-R/06) based upon the United States Food and Drug Administration Redbook Guidelines for Reproduction Studies IV.C.9.a and Developmental Toxicity Studies IV.C.9.b.
Deviations:
no
GLP compliance:
yes
Limit test:
no

Test material

Constituent 1
Reference substance name:
Chromium(3+) tri(pyridine-3-carboxylate)
Cas Number:
64452-96-6
Molecular formula:
Cr(C6H4NO2)3
IUPAC Name:
Chromium(3+) tri(pyridine-3-carboxylate)
Test material form:
solid: particulate/powder
Specific details on test material used for the study:
Niacin bound chromium (NBC) is a unique, patented oxygen-coordinated niacin-bound chromium complex commercially known as ChromeMate CM-100M (powder) and was obtained from InterHealth Nutraceuticals, Benicia, CA, USA. Lot#306013

Test animals

Species:
rat
Strain:
Sprague-Dawley
Sex:
male/female
Details on test animals or test system and environmental conditions:
TEST ANIMALS
- Source: bred and reared at the animal breeding facility of INTOX Pvt. Ltd. India
- Age at study initiation: 7–9 weeks old
- Weight at study initiation: F0: 263 ± 18g , F1: 245 ± 17g
- Diet: ad libitum, ‘‘Nutrilab” brand extruded rodent powdered feed manufactured by M/s Vetcare Pvt. Ltd., Bangalore, India, and tested for nutrients and contaminants, was provided ad libitum to the animals during the study period.
- Water: ad libitum
- Acclimation period: The animals were allowed to acclimatize at least one week before the initiation of experiments

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 19 and 25 °C
- Humidity (%): 30–70%
- Air changes (per hr): 10–15 air changes per h.
- Photoperiod (hrs dark / hrs light): 12 h light/dark cycle

Administration / exposure

Route of administration:
oral: feed
Vehicle:
unchanged (no vehicle)
Details on exposure:
DIET PREPARATION
- Rate of preparation of diet (frequency): The experimental diets were prepared once a week.
- Mixing appropriate amounts with (Type of food): NBC was mixed with powdered rodent diet (Nutrilab” brand extruded rodent powdered feed manufactured by M/s Vetcare Pvt. Ltd., Bangalore, India) to obtain the three concentration levels. Initially, a small volume of dietpremix was prepared which was then mixed with remaining portion of diet in a mechanized ribbon blender for about 20 min to obtain desired homogeneity of the test article concentration in diet.
Details on mating procedure:
The male and female rats of F0 generation from each dose group were mated and allowed to deliver normally. No further details on mating procedure was provided in the article.
Analytical verification of doses or concentrations:
no
Duration of treatment / exposure:
The treatment groups of the F0 parental generation: Receiving feed containing 4, 15, and 60 ppm NBC for a period of 10 weeks before mating, throughout mating, and continued until their termination. The exposure was continued through the next generations, up to completion of developmental toxicity study. At weaning, one male and one female pup from each litter from control and treatment dose groups were selected for first filial (F1 ) generation. The selected F1 animals were exposed to NBC for 10 weeks before mating and then they were mated to produce second generation (F2a).
Frequency of treatment:
continuously via food
Details on study schedule:
- F1 parental animals not mated until 10 weeks after selected from the F1 litters.
- Age at mating of the mated animals in the study: 10 weeks
Doses / concentrationsopen allclose all
Dose / conc.:
60 ppm
Remarks:
nominal in diet
Dose / conc.:
15 ppm
Remarks:
nominal in diet
Dose / conc.:
4 ppm
Remarks:
nominal in diet
Dose / conc.:
0 ppm
Remarks:
nominal in diet
No. of animals per sex per dose:
30 males and 30 females per dose group
Control animals:
yes, plain diet
Details on study design:
- Dose selection rationale: Since NBC is intended to be consumed by human beings up to a maximum dose of 4 mg of NBC/day, the highest dose level for this study was selected so as not to exceed 100 times the maximum recommended human dose, which has a dietary equivalent concentration of 60 ppm. A dose range study revealed no adverse effects of NBC on body weight, feed consumption, mating behavior, fertility, gestation or lactation in rat at dose level up to 60 ppm.
- Rationale for animal assignment (if not random): Sprague–Dawley rats (30/group/sex) were randomly divided into one control and three treatment groups (low, mid and high).
Positive control:
none required

Examinations

Parental animals: Observations and examinations:
CAGE SIDE OBSERVATIONS: Yes
- Time schedule: Animals were examined daily for signs of toxicity and mortality during the entire period of the study.

DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: clinical observations recorded for the parental male and female rats of both (F0 and F1) the generations during the premating and mating periods

BODY WEIGHT: Yes
- Time schedule for examinations: Body weight and feed consumption of the animals from each group were recorded 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, feed consumption was calculated as g/rat/day

WATER CONSUMPTION AND COMPOUND INTAKE (if drinking water study): No data

Oestrous cyclicity (parental animals):
For both F0 and F1 parental animals reproductive parameters such as estrous cycle, female fertility index, gestation index, live-born index, mean litter size, sex ratio (at birth), number of stillbirths at day 0, number of live births at day 0, survival index were assessed.
Sperm parameters (parental animals):
For both F0 and F1 parental animals reproductive parameters such as sperm count (epididymal and homogenization resistant testicular), sperm motility and sperm morphology were assessed.
Litter observations:
STANDARDISATION OF LITTERS
- Performed on day 4 postpartum: no

PARAMETERS EXAMINED
The following parameters were examined in [F1 / F2] offspring: clinical observations and mortality during lactation, litter observations, sexual maturation, physical development endpoints(such as unfolding of pinna (UP), hair growth (HG), teeth eruption (TE), eye opening (EYO) and ear opening (EO)), organ weights, necropsy and histopathology changes

GROSS EXAMINATION OF DEAD PUPS: Yes, all pups, not selected for next generation, were sacrificed on lactation day 21, and subjected to necropsy.
Postmortem examinations (parental animals):
SACRIFICE
- Male animals: After the mating and following confirmation of pregnancy status, males were euthanized with carbon dioxide and were subjected to necropsy and evaluation of sperm parameters such as sperm motility, cauda epididymis sperm count, testicular spermatid head count and sperm morphology.
- Maternal animals: All females were killed after weaning.

GROSS NECROPSY
- At necropsy after the mating period (male rats) or the lactation period (female rats) the group mean values of absolute and relative weight (% of body weight and % of brain weight) of liver, kidneys, brain, spleen, adrenals, pituitary, testes, seminal vesicles (with cowper’s glands), prostate, epididymides, ovaries and uterus, of male and/or female parental rats of F0 generation and F1 generation exposed to NBC at levels of 4, 15, and 60 ppm did not reveal any significant differences from the respective control group,

HISTOPATHOLOGY / ORGAN WEIGHTS
Organ weights were recorded at necropsy, and weighed organs from 10 randomly selected animals were subjected to histopathological examination.
Postmortem examinations (offspring):
SACRIFICE
- The F1 offspring not selected as parental animals and all F2 offspring were sacrificed at : All pups, not selected for next generation, were sacrificed on lactation day 21, and subjected to necropsy.

GROSS NECROPSY
- Gross necropsy consisted of brain, thymus and spleen

HISTOPATHOLOGY / ORGAN WEIGTHS: Histological examination of brain, thymus and spleen of pups euthanized at the end of lactation period
Statistics:
For statistical analysis, a litter was considered as the basic sampling unit. To analyze the various parameters, different and appropriate statistical methods were employed. For data on parental body weight and weight gain, feed intake, and organ weights Bartlett’s test followed by ANOVA and Dunnett’s test was employed with statistical significance at P < 0.05. Day 0 and absolute body weights were analyzed by paired t-test. Group differences in litter size were analyzed by Student t-test. Sex ratio was assessed by Chi-square test (2 x 2 contingency tables). Results of the statistical analysis were described as significantly higher (+)/lower (-) than control values at P < 0.05.

Results and discussion

Results: P0 (first parental generation)

General toxicity (P0)

Clinical signs:
effects observed, non-treatment-related
Description (incidence and severity):
All deaths and abnormal clinical signs observed in the rats during F0 and F1 generations, such as transient/ reversible spells of emaciation, abdominal breathing, respiratory rales, hypoactivity, circling disorder and lacrimation, were considered to be incidental and not due to NBC feeding.
Dermal irritation (if dermal study):
not examined
Mortality:
mortality observed, non-treatment-related
Description (incidence):
All deaths and abnormal clinical signs observed in the rats during F0 and F1 generations, such as transient/ reversible spells of emaciation, abdominal breathing, respiratory rales, hypoactivity, circling disorder and lacrimation, were considered to be incidental and not due to NBC feeding.
Body weight and weight changes:
no effects observed
Food consumption and compound intake (if feeding study):
no effects observed
Food efficiency:
not specified
Water consumption and compound intake (if drinking water study):
not examined
Ophthalmological findings:
not specified
Haematological findings:
not specified
Clinical biochemistry findings:
not specified
Endocrine findings:
not specified
Urinalysis findings:
not examined
Behaviour (functional findings):
not examined
Immunological findings:
not examined
Organ weight findings including organ / body weight ratios:
no effects observed
Histopathological findings: non-neoplastic:
no effects observed
Histopathological findings: neoplastic:
not specified
Other effects:
no effects observed

Reproductive function / performance (P0)

Reproductive function: oestrous cycle:
no effects observed
Description (incidence and severity):
Similarly, the unaltered length and normalization of estrous cycles in treated females, mating performance as evidenced from unaltered indices of male fertility and female fertility, maintenance of normal gestation was evident from unaltered gestation length and gestation indices.
Reproductive function: sperm measures:
no effects observed
Description (incidence and severity):
Sperm evaluation. For both the F0 and F1 generations exposed to NBC at dose level up to 60 ppm, evaluations of sperm parameters of male rats during premating and mating period, and the period thereafter up to their termination, did not reveal any changes that could be attributed to the test article
Reproductive performance:
no effects observed
Description (incidence and severity):
NBC at dose levels up to 60 ppm during premating and mating periods and gestation period in females did not reveal any treatment related adverse effects on reproductive performance in terms of fertility and mating, gestation

Details on results (P0)

1. Parental observations (F0 and F1 )
1.1. Food consumption and test compound dosage determination Data on feed consumption by the parental male and female rats of both (F0 and F1 ) the generations during the premating and mating periods, for both sexes, and during gestation and lactation in case of female rats, did not reveal any significant treatment related changes in the average daily feed intake by the male and female rats compared to the respective control groups, across the different dose levels for each of the F0 and F1 generations and also when compared across these two generations. Based on feed intake, the resulting dose of NBC during premating period for the highest dose groups F0 male and female was calculated as 5.88 and 8.24 mg/kg/day, respectively, for F0 generation, while the same was, respectively, 9.71 and 9.83 mg/kg/day in case of F1 generation. The total daily dose of NBC for all groups is presented in Table 2.
1.2. Clinical signs and mortality
Compared to the respective control groups, across the different dose levels for each of the F0 and F1 generations and also when compared across these two generations, results of survival and clinical observations recorded for the parental male and female rats of both (F0 and F1 ) the generations during the premating and mating periods, for both sexes, and during gestation and lactation in case of female rats, did not reveal any remarkable incidence of mortality and abnormal clinical signs among the male and female rats exposed to NBC. All deaths and abnormal clinical signs observed in the rats during F0 and F1 generations, such as transient/reversible spells of emaciation, abdominal breathing, respiratory rales, hypoactivity, circling disorder and lacrimation, were considered to be incidental and not due to NBC feeding.
1.3. Body weights
The average body weight and body weight gains of the parental male and female rats of both the generations (F0 and F1 ) during the premating and mating periods, and during gestation and lactation of female rats, did not reveal any remarkable alterations which could be attributed to NBC exposure at any of the doses, when compared to the respective control groups, across the different dose levels for each of the F0 and F1 generations and also when compared across these two generations. Although, other occasional instances of group mean values of treated animals differing from those of the respective control groups were noted, these were considered incidental or of no toxicological significance due either their lack of dose relation, their small magnitudes, or other procedural reasons unrelated to the treatment.
1.4. Mating, fertility and reproduction
Exposure of male and female rats, from both the F0 and F1 generations, to NBC at dose levels up to 60 ppm during premating and mating periods and gestation period in females did not reveal any treatment related adverse effects on reproductive performance in terms of fertility and mating, gestation, parturition and the litters born. Similarly, the unaltered length and normalization of estrous cycles in treated females, mating performance as evidenced from unaltered indices of male fertility and female fertility, maintenance of normal gestation was evident from unaltered gestation length and gestation indices. Furthermore the pups born alive were unaffected, as evidenced from their live birth indices and did not reveal any treatment related adverse effects.
The values of male fertility indices for treatment groups in F0 and F1 generations did not differ significantly from those of the controls, and also compared well with the historical control data at the test facility. The values of male fertility indices for treatment groups in F0 generation in control, 4, 15 and 60 ppm groups were 90%, 100%, 80% and 92.2%, respectively. Similarly, these values for treatment groups in F1 generation were 103%, 100% and 100% at the doses of 4, 15, and 60 ppm, respectively, while the value was 96% for the control group, respectively.
1.4.1. Sperm evaluation. For both the F0 and F1 generations exposed to NBC at dose level up to 60 ppm, evaluations of sperm parameters of male rats during premating and mating period, and the period thereafter up to their termination, did not reveal any changes that could be attributed to the test article. This was evident by virtue of the group mean values of motility of sperms in cauda epididymis, counts of sperms in cauda epididymis (absolute count and per gram of cauda weight), counts of homogenization resistant spermatids (absolute count and per gram of testis weight) per testis, and the morphological evaluations of the sperms by microscopy of stained smears. Although the sperm motility of F1 parents compared to the F0 parents was found to be slightly lower, it was not considered to be related to the NBC exposure, as the lowering was also observed in the concurrent control group of rats and the altered values were comparable to the historical control data.
1.5. Parental organ weights, necropsy and histopathology: At necropsy after the mating period (male rats) or the lactation period (female rats) the group mean values of absolute and relative weight (% of body weight and % of brain weight) of liver, kidneys, brain, spleen, adrenals, pituitary, testes, seminal vesicles (with cowper’s glands), prostate, epididymides, ovaries and uterus, of male and/or female parental rats of F0 generation and F1 generation exposed to NBC at levels of 4, 15, and 60 ppm did not reveal any significant differences from the respective control group, which could be ascribed to NBC. Necropsy and histological examinations performed on the parents of the F0 and F1 generations, which died during the study or were terminated at end of the mating period (males) or the lactation period (females), did not reveal any incidence of gross and microscopic pathological alterations attributable to their exposure to NBC at dose levels of up to 60 ppm. All the gross and microscopic findings noted were considered to be incidental as the incidence was found to be comparable among the control group and the treatment groups, without any dose dependent trend.

Effect levels (P0)

open allclose all
Dose descriptor:
NOAEL
Effect level:
60 ppm
Based on:
test mat.
Remarks:
The equivalent dose in male and female rats was 7.80 and 8.31 mg/kg/day in male and female rats, respectively.
Sex:
male/female
Basis for effect level:
other: no adverse effects observed
Remarks on result:
other: The parental and offspring no-observed-adverse-effect level (NOAEL) in this two-generation reproduction toxicity study was found to exceed 60 ppm in diet.
Dose descriptor:
NOAEL
Effect level:
7.8 mg/kg bw/day
Based on:
test mat.
Sex:
male
Basis for effect level:
other: no adverse effects observed
Dose descriptor:
NOAEL
Effect level:
8.31 mg/kg bw/day
Based on:
test mat.
Sex:
female
Basis for effect level:
other: no adverse effects observed

Target system / organ toxicity (P0)

Critical effects observed:
no

Results: P1 (second parental generation)

General toxicity (P1)

Clinical signs:
effects observed, non-treatment-related
Description (incidence and severity):
All deaths and abnormal clinical signs observed in the rats during F0 and F1 generations, such as transient/ reversible spells of emaciation, abdominal breathing, respiratory rales, hypoactivity, circling disorder and lacrimation, were considered to be incidental and not due to NBC feeding.
Dermal irritation (if dermal study):
not specified
Mortality:
mortality observed, non-treatment-related
Description (incidence):
All deaths and abnormal clinical signs observed in the rats during F0 and F1 generations, such as transient/ reversible spells of emaciation, abdominal breathing, respiratory rales, hypoactivity, circling disorder and lacrimation, were considered to be incidental and not due to NBC feeding.
Body weight and weight changes:
no effects observed
Food consumption and compound intake (if feeding study):
no effects observed
Food efficiency:
not specified
Water consumption and compound intake (if drinking water study):
not examined
Ophthalmological findings:
not specified
Haematological findings:
not specified
Clinical biochemistry findings:
not specified
Endocrine findings:
not specified
Urinalysis findings:
not specified
Behaviour (functional findings):
not specified
Immunological findings:
not specified
Organ weight findings including organ / body weight ratios:
no effects observed
Gross pathological findings:
no effects observed
Neuropathological findings:
not specified
Histopathological findings: non-neoplastic:
no effects observed
Histopathological findings: neoplastic:
not specified
Other effects:
no effects observed

Reproductive function / performance (P1)

Reproductive function: oestrous cycle:
no effects observed
Description (incidence and severity):
Similarly, the unaltered length and normalization of estrous cycles in treated females, mating performance as evidenced from unaltered indices of male fertility and female fertility, maintenance of normal gestation was evident from unaltered gestation length and gestation indices.
Reproductive function: sperm measures:
no effects observed
Description (incidence and severity):
Sperm evaluation. For both the F0 and F1 generations exposed to NBC at dose level up to 60 ppm, evaluations of sperm parameters of male rats during premating and mating period, and the period thereafter up to their termination, did not reveal
any changes that could be attributed to the test article
Reproductive performance:
no effects observed
Description (incidence and severity):
NBC at dose levels up to 60 ppm during premating and mating periods and gestation period in females did not reveal any treatment related adverse effects on reproductive performance in terms of fertility and mating, gestation
The values of male fertility indices for treatment groups in F0 and F1 generations did not differ significantly from those of the controls, and also compared well with the historical control data at the test facility

Effect levels (P1)

Dose descriptor:
NOAEL
Effect level:
60 ppm
Based on:
test mat.
Remarks:
The equivalent dose in male and female rats was 7.80 and 8.31 mg/kg/day in male and female rats, respectively.
Sex:
male/female
Basis for effect level:
other: no adverse effects observed
Remarks on result:
other: The parental and offspring no-observed-adverse-effect level (NOAEL) in this two-generation reproduction toxicity study was found to exceed 60 ppm in diet.

Target system / organ toxicity (P1)

Critical effects observed:
no

Results: F1 generation

General toxicity (F1)

Clinical signs:
no effects observed
Dermal irritation (if dermal study):
not examined
Mortality / viability:
no mortality observed
Body weight and weight changes:
no effects observed
Food consumption and compound intake (if feeding study):
no effects observed
Food efficiency:
not specified
Water consumption and compound intake (if drinking water study):
not specified
Ophthalmological findings:
not specified
Haematological findings:
not specified
Clinical biochemistry findings:
not specified
Urinalysis findings:
not examined
Sexual maturation:
no effects observed
Anogenital distance (AGD):
not specified
Nipple retention in male pups:
not specified
Organ weight findings including organ / body weight ratios:
no effects observed
Gross pathological findings:
no effects observed
Histopathological findings:
no effects observed
Other effects:
not specified

Developmental neurotoxicity (F1)

Behaviour (functional findings):
not specified

Developmental immunotoxicity (F1)

Developmental immunotoxicity:
not specified

Details on results (F1)

2. Offspring observations (F1 and F 2a )
2.1. Offspring body weights: The data on average values of body weights of offspring of both the generations (F1 and F 2a ) recorded on lactation days 0, 4, 7, 14 and 21, did not reveal any alterations which could be attributed to exposure of their dams to NBC at levels up to 60 ppm, when compared to the respective control groups, across the different dose levels for each of the F0 and F1 generations and also when compared across these two generations.
2.2. Offspring clinical observations and mortality during lactation: Compared to the respective control groups of pups, across the different dose levels for each of the F1 and F2a generations and also when compared across two generations, data on survival and clinical observations recorded for the offspring of both the generations (F1 and F2a ) during lactation period of 21 days did not reveal any remarkable differences. The observations included clinical abnormalities in pups, and the incidence of normal pups, pups found dead on lactation day 0 and thereafter, pups cannibalized by the dam on lactation day 0 and thereafter, and pups which were terminated in moribund state.
2.3. Litter observations: Comparison of the offspring data with respective control groups and also across the F1 and F2a generations did not reveal any adverse effect on their litter sizes, the sex ratios of litters, the live birth indices and the viability indices of litters calculated for days 4, 7, 14 and 21 of lactation, following exposure of parental females to NBC at dose levels of 4, 15, and 60 ppm.
2.4. Offspring sexual maturation: The sexual maturation was measured only for F2a generation, in terms of age at which there is balano-preputial separation in males and vaginal opening in females. Exposure to NBC at any of the dose levels did not affect the age of sexual maturity by the offspring belonging to the F2a generation. The group mean age at balano-preputial separation in male pups was 31.4 ± 2.58 days, 30.2 ± 2.76 days, 28.9 ± 1.80 days and 27.7 ± 1.42 days, respectively, for the control group and low, mid and high dose levels.
Historical control value (Mean + SD) for the same was 24.6 ± 2.7. The group mean age at vaginal opening in female pups was 57.4 ± 6.85 days, 54.4 ± 8.64 days, 51.7 ± 9.06 days and 50.8 ± 9.64 days, respectively, for the control group and low, mid and high dose levels. Historical control value (Mean + SD) for the same was 49.7 ± 5.3. Although there was a trend for an inverse relationship between dose and sexual maturation in F2a generation, this trend was not statistically significant.
2.5. Offspring physical development endpoints: Exposure of the parental animals of both the F0 and F1 generation to NBC at 4, 15 and 60 ppm had no significant (P > 0.05) adverse effect on the physical development of their litters during the period of lactation, which was evident by the unaltered period, in days, required by pups of F1 and F2a generation to attain certain landmarks of physical development such as days required for unfolding of ear pinna, hair growth on the body, time (days) for eruption of teeth, opening of eyes and opening of ear, compared to the respective control groups.
2.6. Offspring organ weights: Compared to the respective control group, exposure of the parental animals of both the F0 and F1 generation to NBC at dose levels of up to 60 ppm did not affect the organ weights of their F1 and F2a offspring. The group mean values of absolute and relative weights (% of body weights and% of brain weights) of brain, spleen and thymus of pups of F1 and F2a generation did not significantly alter between the control and treatment groups (Table 8).
2.7. Offspring necropsy and histopathology changes: Necropsy performed on the offspring of the F1 and F2a generations, on day 4 or at end of the lactation period, and histological examination of brain, thymus and spleen of pups euthanized at the end of lactation period did not reveal any incidence of gross or microscopic pathological alterations attributable to exposure of their parents to NBC at the dose levels of up to 60 ppm. All the gross and microscopic pathology findings encountered in this study were considered incidental as the incidence was found to be comparable among the control group and the treatment groups, without any dose dependent trend. Thus, NBC treatment did not cause any significant histopathological changes in any organ.

Effect levels (F1)

Dose descriptor:
NOAEL
Generation:
F1
Effect level:
60 ppm
Based on:
test mat.
Remarks:
The equivalent dose in male and female rats was 7.80 and 8.31 mg/kg/day in male and female rats, respectively.
Sex:
male/female
Basis for effect level:
other: no adverse effects observed
Remarks on result:
other: The parental and offspring no-observed-adverse-effect level (NOAEL) in this two-generation reproduction toxicity study was found to exceed 60 ppm in diet.

Target system / organ toxicity (F1)

Critical effects observed:
no

Results: F2 generation

General toxicity (F2)

Clinical signs:
no effects observed
Dermal irritation (if dermal study):
not examined
Mortality / viability:
no mortality observed
Body weight and weight changes:
no effects observed
Food consumption and compound intake (if feeding study):
not specified
Food efficiency:
not specified
Water consumption and compound intake (if drinking water study):
not specified
Ophthalmological findings:
not specified
Haematological findings:
not specified
Clinical biochemistry findings:
not specified
Urinalysis findings:
not specified
Sexual maturation:
no effects observed
Anogenital distance (AGD):
not specified
Nipple retention in male pups:
not specified
Organ weight findings including organ / body weight ratios:
no effects observed
Gross pathological findings:
no effects observed
Histopathological findings:
no effects observed
Other effects:
no effects observed

Developmental neurotoxicity (F2)

Behaviour (functional findings):
not specified

Developmental immunotoxicity (F2)

Developmental immunotoxicity:
not specified

Effect levels (F2)

Dose descriptor:
NOAEL
Generation:
F2a
Effect level:
60 ppm
Based on:
test mat.
Remarks:
The equivalent dose in male and female rats was 7.80 and 8.31 mg/kg/day in male and female rats, respectively.
Sex:
male/female
Basis for effect level:
other: no adverse effects observed
Remarks on result:
other: The parental and offspring no-observed-adverse-effect level (NOAEL) in this two-generation reproduction toxicity study was found to exceed 60 ppm in diet.

Target system / organ toxicity (F2)

Critical effects observed:
no

Overall reproductive toxicity

Reproductive effects observed:
not specified

Applicant's summary and conclusion

Conclusions:
In this two generation reproduction toxicity study with Niacin bound chromium (NBC) no adverse or non-adverse effects to rats were noted up to the maximum dose tested (60 ppm in diet, equivalent to 8 mg/kg bw/d) and thus the NOAEL = NOEL can be set to the highest dose tested in this study being 8 mg/kg bw/d. No effects on fertility, developmental toxicity or through lactation were seen in rats.
Executive summary:

The findings of this two-generation reproduction toxicity study demonstrate that exposure of male and female Sprague–Dawley rats to Niacin bound chromium (NBC) at the dietary dose levels of 4, 15, and 60 ppm, (approximately corresponding to 0.5, 2 and 8 mg/kg bw/day, respectively) for over two generations was without any adverse effects on various parameters of reproductive performance such as growth, sexual maturity, fertility and mating, gestation, parturition, litter properties, lactation and development of their offspring. NBC, at these dose levels, did not induce any systemic toxicity in the parental rats and their offspring. The present two-generation study serves as a better model for observing the influences of NBC on germ cell development, spermatogenesis, and sexual maturity. Results from the present study did not reveal any adverse effects of NBC on spermatogenesis at dose levels up to 60 ppm in F0 and F1 generation of male rats, respectively.

Overall, the results of two-generation reproductive toxicity in conjunction with earlier studies suggest that under the conditions of the study NBC is safe in male and female rats. The parental and offspring no-observed-adverse-effect level (NOAEL) in this two-generation reproduction toxicity study was found to exceed 60 ppm in diet. The equivalent dose in male and female rats was 7.80 and 8.31 mg/kg/day in male and female rats, respectively.

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