Nickel bis(sulphamidate)

Repeated dose toxicity: oral

Administrative data

Endpoint:

repeated dose toxicity: oral, other

Remarks:

combined repeated dose and carcinogenicity

Type of information:

experimental study

Adequacy of study:

key study

Study period:

not reported

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Cross-reference

Reason / purpose for cross-reference:

reference to same study

Data source

Reference

Reference Type:

publication

Title:

Unnamed

Year:

2007

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

other: OECD 451 (Carcinogenicity Studies)

Test guideline 2

Qualifier:

according to guideline

Guideline:

other: EPA OPPTS 870.4200 (Carcinogenicity)

GLP compliance:

yes

Limit test:

no

Test material

Test material information

1

Reference substance name:

Nickel sulphate

EC Number:

232-104-9

EC Name:

Nickel sulphate

Cas Number:

7786-81-4

Molecular formula:

NiSO4

IUPAC Name:

nickel(2+) sulfate

Constituent 1

Reference substance name:

Nickel sulfate hexahydrate

Cas Number:

10101-97-0

Molecular formula:

NiSO4.6H2O

IUPAC Name:

Nickel sulfate hexahydrate

Test animals

Species:

rat

Strain:

Fischer 344

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Charles River Laboratories, Inc., Raleigh, North Carolina.
- Age at study initiation: 6 weeks
- Weight at study initiation: 118 to 147 g for males and 93 to 112 g for females
- Fasting period before study: not reported
- Housing: housed individually in suspended stainless steel cages that were rotated in a regular fashion
- Diet (e.g. ad libitum): ad libitum
- Water (e.g. ad libitum): ad libitum
- Acclimation period: acclimated to the laboratory conditions prior to in-life initiation
- Other: The results of the pretest health screen (gross necropsy and serological analyses) conducted prior to in-life initiation indicated that the population of animals was suitable for study use. Serological analyses of blood samples fromfive male and five female sentinel animals conducted by
BioReliance Corporation, Rockville, Maryland, during weeks 25, 51, 77 and 103 did not reveal the presence of any viral infections that would
negatively impact the results of this study.

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 70 to 76 °F
- Humidity (%): 29 to 73%
- Air changes (per hr): 10 to 15 air changes per hour
- Photoperiod (hrs dark / hrs light): 12-h light/12-h dark cycle

IN-LIFE DATES: not reported

Administration / exposure

Route of administration:

oral: gavage

Vehicle:

water

Details on oral exposure:

PREPARATION OF DOSING SOLUTIONS: a specified amount of the test article and vehicle was mixed weekly. The mixtures were stirred continuously throughout each exposure period. The appearance of each test article preparation was determined and documented as a clear colorless solution for groups 2 and 3 (10 and 30mg/kg/day) and a clear pale blue solution for group 4 (50 mg/kg/day).

DIET PREPARATION
- not applicable

VEHICLE
- water

Analytical verification of doses or concentrations:

yes

Details on analytical verification of doses or concentrations:

Analyses were conducted by KAR Laboratories, Inc. (Kalamazoo, Michigan) prior to study initiation, during week 51, and following study completion
to confirm the stability and purity of the test substance. Reverse osmosis deionized tap water was used for administration to control animals and in
the preparation of the test article mixtures. Analytical concentration verification analyses conducted throughout the study demonstrated that the
exposure solutions were stable and properly prepared. All analyses were within ±10% of the nominal concentration.

Duration of treatment / exposure:

104 weeks

Frequency of treatment:

daily

Doses / concentrations

Remarks:

Doses / Concentrations:
10, 30 and 50 mg/kg/day
Basis:
other: dose via oral gavage

No. of animals per sex per dose:

60 males/60 females per dose

Control animals:

yes, concurrent no treatment

Details on study design:

- Dose selection rationale: Exposures for the 90-day range finding study were selected based on previous gavage studies of nickel sulfate hexahydrate in rats. The 90-day range-finding study of nickel sulfate hexahydrate administered by gavage was conducted using exposures of 0, 50, 75, 100, 125, and 150 mg/kg. Findings from this study are reported in the Results section. Based on the data from the 90-day range-finding study, exposure levels of 10, 30 and 50 mg/kg/day were selected for the 2 year oral gavage carcinogenicity study.

- Rationale for animal assignment (if not random): Sixty female and sixty male animals were assigned to each exposure group using a computer randomization program.

Positive control:

None reported

Examinations

Observations and examinations performed and frequency:

CAGE SIDE OBSERVATIONS: Yes
- Time schedule: General health/mortality/moribundity checks were performed twice daily.

DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: Detailed clinical observations were performed weekly and on the day of scheduled euthanasia (weeks 104–105). Beginning on week 25, detailed clinical observations included a palpable mass examination (including the occurrence, size, location and description of any palpable masses) followed by persistence or disappearance of these masses being documented at the next weekly clinical observation.

BODY WEIGHT: Yes
- Time schedule for examinations: Individual body weights were recorded prior to randomization (day −3), on day 0 (i.e., the start of exposure), weekly during the first 13 weeks, once every 4 weeks thereafter and during week 103.

FOOD CONSUMPTION AND COMPOUND INTAKE (if feeding study):
- Individual food consumption (grams/animal/day) was recorded on day 0, weekly during the first 13 weeks and once every 4 weeks thereafter, with the final food consumption measurement during week 103.

HAEMATOLOGY: Yes
- Selected hematological parameters were measured in blood samples collected from 10 animals/sex/group during week 54 (via tail vein) and prior to scheduled euthanasia during week 104/105 (via orbital plexus). Hematology and clinical chemistry parameters were measured according to the OECD 451 protocol.

Sacrifice and pathology:

GROSS PATHOLOGY/HISTOPATHOLOGY:
All animals were subjected to a complete gross necropsy examination at the time of death or euthanasia. Tissues collected at necropsy from all
animals were processed for histopathological evaluation. Slides were prepared by Histo Techniques (Powell, Ohio) and Charles River Laboratories-
Pathology Associates (Frederick, Maryland) and were examined microscopically by a Charles River Laboratories board-certified veterinary
pathologist.

Other examinations:

Near the end of the study (week 103), additional biological sampling was performed to provide data on nickel in urine, feces and blood. Immediately
following exposure on 1 day during week 103, five females and five males from each exposure group were placed in urine collection cages equipped with fecal collection screens, and an ice bath for cooling collected urine samples. Blood was collected from the orbital plexus of each animal
approximately 30 min and 24 h post-exposure and sent to WIL Research Laboratories, Inc. (Ashland, Ohio) for analysis of blood nickel
concentration. Urine and fecal samples were collected from each cage approximately 24 h post-exposure and sent to KAR Laboratories, Inc. for
urine and fecal analysis of nickel concentrations. Urine was analyzed also for creatinine and albumin concentrations. Other standard hematology
and clinical chemistry parameters for blood as well as other standard urinalysis parameters for urine were measured by Charles River Laboratories.

Statistics:

In-life data: The data were initially tested for normality using Levene's test for equality of variance followed by the Shapiro–Wilks test for normality.
A p≤0.001 level of significance was required for either test to reject the assumptions. If both assumptions were fulfilled, a singlefactor
ANOVA was applied, with animal grouping as the factor, utilizing a p≤0.05 level of significance. If the parametric ANOVA was significant at p≤0.05,
Dunnett's test was used to identify statistically significant differences between the control group and each nickel sulfate-treated group at the 0.05,
0.01 and 0.001 levels of significance. If either of the parametric assumptions was not satisfied, then the Kruskal–Wallis nonparametric ANOVA
procedure was used to evaluate intergroup differences (p≤0.05). The Dunn's multiple comparison test was applied if this ANOVA was significant,
again utilizing significance levels of p≤0.05, 0.01 and 0.001.

Survival Data: Kaplan–Meier estimates of group survival rates were calculated, by sex, and shown graphically. A log-rank dose response trend test
of survival rates was performed utilizing dose coefficients. In addition, a log-rank test for survival was used to make pairwise comparisons of each
treated group with the control group. Both the trend test and pairwise comparisons were conducted at the 0.05 significance level.

Results and discussion

Results of examinations

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):

no effects observed

Ophthalmological findings:

not examined

Haematological findings:

no effects observed

Clinical biochemistry findings:

no effects observed

Urinalysis findings:

no effects observed

Behaviour (functional findings):

not examined

Organ weight findings including organ / body weight ratios:

not examined

Gross pathological findings:

effects observed, treatment-related

Histopathological findings: non-neoplastic:

effects observed, treatment-related

Histopathological findings: neoplastic:

no effects observed

Details on results:

MORTALITY:
Group (Dose) Males (deaths/N, % mortality) Females(deaths/N, % mortality)
1 (0 mg/kg/day) 36/60 (60) 14/60 (23)
2 (10 mg/kg/day) 29/60 (48) 20/60 (33)
3 (30 mg/kg/day) 30/60 (50) 26/60 (43)
4 (50 mg/kg/day) 34/60 (57) 27/60 (45)

BODY WEIGHT AND WEIGHT GAIN:
Body weights decreased in a exposure-dependent manner, with significantly decreased body weights observed in the two highest exposure groups for males and females. Reductions in weight gain relative to controls at study week 103 reached the level of biological significance (i.e., >10% decrease) in the group 3 and 4 males and the group 4 females. This significant weight reduction indicates that the Maximum Tolerated Dose was reached in this study for both males and females.

HAEMATOLOGY:
A few statistically significant differences in the hematology data were observed in the nickel sulfate-treated males and females. However, none of these differences was suggestive of a hyperplastic (i.e., leukemia) response and none of these changes was considered toxicologically meaningful since they were small and did not follow a consistent exposure-related pattern.

GROSS PATHOLOGY/HISTOPATHOLOGY:
Gross necropsy and histopathology observations: Numerous gross necropsy findings were observed for animals in the control and nickel
sulfate-treated groups but the type and incidence of these findings observed for the treated animals were comparable to those observed in the
control group, and were consistent with findings commonly seen in aging rats in a longterm study. None of the neoplastic or non-neoplastic
microscopic findings was considered to be related to the experimental exposures. The non-neoplastic findings were either considered to
be secondary to toxicity or incidental background occurrences rather than a direct effect of nickel sulfate.

The pathology report, pathology peer-review and the pathology working group concurred that nickel sulfate hexahydrate did not
cause any carcinogenic effects in this study. Analysis of the tumor data revealed only one statistically significant (p<0.001) increase in tumors
corresponding to keratoacanthoma (tail) in the group 2 males. However, this finding is of questionable toxicologic significance since there was no
exposure–response relationship, the incidence rate in the group 2 males (15%) was only slightly higher than the upper end of Haseman's historical
control incidence for this tumor type (0–14%) and the incidence rate in the remaining control and treated groups (0–7%) was within the range of the
CRL-Ohio historical incidence (0–2%) and the Haseman historical incidence (0–14%). No other tumor finding in this study was statistically significant.

No notable differences were observed between controls and treated animals for the hematology, biochemistry and urinalysis parameters measured
during the toxicokinetic satellite study.

Effect levels

1

Dose descriptor:

NOAEL

Effect level:

2.2 other: mg Ni/kg bw/day

Sex:

male/female

Basis for effect level:

other: significant decrease in body weight

2

Dose descriptor:

LOAEL

Effect level:

6.7 other: mg Ni/kg bw/day

Sex:

male/female

Basis for effect level:

other: significant decrease in body weight

Target system / organ toxicity

Critical effects observed:

not specified