Reaction mass of aluminium fluoride and chromium trifluoride and nickel difluoride

Repeated dose toxicity: inhalation

Administrative data

Endpoint:

chronic toxicity: inhalation

Type of information:

migrated information: read-across from supporting substance (structural analogue or surrogate)

Adequacy of study:

key study

Study period:

September 1985 - June 1990

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: GLP-compliant (13-week and 2-year phases) study, comparable to guideline study, performed by US National Toxicology Program

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

1996

Report date:

1996

Materials and methods

Test guideline

Test guideline 1

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 413 (Subchronic Inhalation Toxicity: 90-Day Study)

Test guideline 2

Qualifier:

equivalent or similar to guideline

Guideline:

other: OECD Guideline 451 (Carcinogenicity studies)

GLP compliance:

yes

Remarks:

(13-week and 2-year phases)

Limit test:

no

Test material

Test material information

Constituent 1

Reference substance name:

Nickel sulfate hexahydrate

IUPAC Name:

Nickel sulfate hexahydrate

Details on test material:

- Name of test material (as cited in study report): Nickel sulfate hexahydrate
- Molecular formula (if other than submission substance): NiSO4.6H2O
- Physical state: Blue-green crystalline powder
- Analytical purity: >98%
- Impurities (identity and concentrations): cobalt (approx. 1500 ppm), silicon (470 ppm), magnesium (120 ppm)
- Lot/batch No.: M062883
- Storage condition of test material: In amber glass bottles at room temperature
- Other: Periodic monitoring of the bulk chemical was performed by Huffman Laboratories, Inc. (Golden, CO) using elemental analyses for nickel, hydrogen, and sulfur prior to and after all studies and every 4 months during the 2-year studies.

Test animals

Species:

other: rats and mice

Strain:

other: F344/N (rats) and B6C3F1 (mice)

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Frederick Cancer Research Facility, Frederick, MD (16-day studies), Simonsen Laboratories, Gilroy, CA (13-week studies), Taconic Farms, Germantown, NY (2-year studies)
- Age at study initiation: 7 weeks (16-day and 13-week studies), 6 weeks (2-year studies)
- Weight at study initiation (g): 160-167 (16-day male rats), 122-125 (16-day female rats), 131-135 (13-week male rats), 106-112 (13-week female rats), 130-133 (2-year male rats), 108-110 (2-year female rats), 21.9-22.7 (16-day male mice), 18.4-19.1 (16-day female mice), 23.0-24.9 (13-week male mice), 17.8-19.2 (13-week female mice), 23.1-23.5 (2-year male mice), 18.8-19.2 (2-year female mice)
- Fasting period before study: Not specified
- Housing: Individually in stainless steel cage units rotated every 4 exposure days (16-day studies) or once weekly (13-week studies) and changed weekly, or stainless steel wire mesh cage units rotated once weekly (2-year studies)
- Diet: ad libitum, except during exposure period, changed at least once weekly
- Water: ad libitum, checked twice daily
- Acclimation period: approximately 3 weeks (16-day and 13-week studies) or 10 days (2-year studies)

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 17.0-28.2 (16-day studies), 19.1-27.6 (13-week studies), 17.2-29.6 (2-year studies)
- Humidity (%): 13-82 (16-day studies), 43-76 (13-week studies), 8 -99 (2-year studies)
- Air changes (per hr): 9-21 (2-year studies)
- Photoperiod (hrs dark / hrs light): 12 / 12

IN-LIFE DATES: From: 23 September 1985 To: 26 June 1990

Administration / exposure

Route of administration:

inhalation: aerosol

Type of inhalation exposure:

whole body

Vehicle:

air

Remarks on MMAD:

MMAD / GSD: MMAD was similar for all exposure concentrations and ranged from 1.8 to 3.1 µm with a geometric standard deviation ranging from 1.6 to 2.9.

Details on inhalation exposure:

GENERATION OF TEST ATMOSPHERE / CHAMBER DESCRIPTION
- Exposure apparatus: Retec nebulizer (In Tox Products, Albuquerque, NM)
- Method of holding animals in test chamber: Whole-body stainless steel chambers
- Source and rate of air: Generator - 3 to 4.5 L/min flow rate
- Method of conditioning air: Not specified
- System of generating particulates/aerosols: Aqueous solution (62.1 g/L in distilled and deionized water) atomized
- Temperature, humidity, pressure in air chamber: See above
- Air flow rate: 3 to 4.5 L/min flow rate
- Air change rate: 9-21 changes/hour (2-year studies)
- Method of particle size determination: Cascade impactor
- Treatment of exhaust air: Not specified

TEST ATMOSPHERE
- Brief description of analytical method used: thermogravimetric analysis (extent of hydration) / electrothermal atomic absorption spectroscopy (nickel content)
- Samples taken from breathing zone: yes

Analytical verification of doses or concentrations:

yes

Details on analytical verification of doses or concentrations:

In the 13-week studies in rats and mice and the 2-year study in rats, the aerosol concentrations were determined gravimetrically from two 3-hour samples (4.5 L/min flow rate) for the 0.12 and 0.25 mg/m3 exposure chambers and from three 2-hour filter samples (3 L/min flow rate) for the higher concentration exposure chambers during each 6-hour exposure day. In the 2-year study in mice, the aerosol concentrations were monitored by collecting three 2-hour filter samples (3 L/min flow rate) from each exposure chamber. The background concentrations of total suspended particles in the control chambers were monitored each exposure day of the 2-year studies by collecting one 6-hour filter sample. In the 2-year studies, the mean concentrations of total suspended particles were 0.02 +/- 0.01 mg particle/mg3 in the rat control chamber and 0.01 +/- 0.01 mg particle/m3 in the mouse control chamber. Good control of aerosol concentration was maintained. A continuous aerosol monitor (Model RAM-S, GCA, Co., Bedford, MA) was used to monitor the stability of the aerosol concentrations and to determine the need to adjust the aerosol generation system during exposures. The RAM-S was used to monitor each chamber for at least 2 minutes at the beginning, middle, and end of each filter sampling period.

Duration of treatment / exposure:

16 days, 13 weeks or 104 weeks

Frequency of treatment:

6 hours and 8 or 12 minutes (to reach 90% of the target concentration) per day, 5 days per week

Doses / concentrations

Doses / concentrations 1

Remarks:

Doses / Concentrations:
Core studies: 0, 3.5, 7, 15, 30, or 60 mg nickel sulfate hexahydrate/m3 (0, 0.7, 1.4, 3.1, 6.1, or 12.2 mg nickel/m3) / Tissue burden studies: 0, 3.5, 15, or 30 mg/m3 (16-day studies)
Basis:
nominal conc.

Doses / concentrations 2

Remarks:

Doses / Concentrations:
Core studies: 0, 0.12, 0.25, 0.50, 1, or 2 mg nickel sulfate hexahydrate/m3 (0, 0.03, 0.06, 0.11, 0.22, or 0.44 mg nickel/m3) / Tissue burden studies: 0, 0.12, 0.5, or 2 mg/m3 (13-week studies)
Basis:
nominal conc.

Doses / concentrations 3

Remarks:

Doses / Concentrations:
0, 0.12, 0.25, or 0.5 mg nickel sulfate hexahydrate/m3 (0, 0.03, 0.06, or 0.11 mg nickel/m3) (2-year core and tissue burden studies in rats)
Basis:
nominal conc.

Doses / concentrations 4

Remarks:

Doses / Concentrations:
0, 0.25, 0.5, or 1 mg nickel sulfate hexahydrate/m3 (0, 0.06, 0.11, or 0.22 mg nickel/m3) (2-year core and tissue burden studies in mice)
Basis:
nominal conc.

No. of animals per sex per dose:

16-day studies (animals/sex/group): 5 (core studies), 4 or 5 (tissue burden studies); 13-week studies (animals/sex/group): 10 (core studies), 5 or 6 (tissue burden studies); 2-year studies (animals/sex/group): Core studies: 5 (7-month and 15-month interim evaluations), 53-55 rats and 60-62 mice (2-year studies); Tissue burden studies: 5 to 7 rats and 5 mice (7-month interim evaluation), 5 (15-month interim evaluation)

Control animals:

yes, concurrent vehicle

Details on study design:

- Dose selection rationale: 16-day studies were used as range-finding studies for 13-week studies which were used themselves as range-finding studies for 2-year studies
- Rationale for animal assignment: Random
- Rationale for selecting satellite groups: Not applicable
- Post-exposure recovery period in satellite groups: Not applicable

Positive control:

Not included

Examinations

Observations and examinations performed and frequency:

CAGE SIDE OBSERVATIONS: Yes
- Time schedule: twice daily

DETAILED CLINICAL OBSERVATIONS: No data

BODY WEIGHT: Yes
- Time schedule for examinations: initially, on day 5 (16-day studies), weekly for 13 weeks (13-week and 2-year studies), monthly thereafter, and at the end of the studies

FOOD CONSUMPTION:
- Food consumption for each animal determined and mean daily diet consumption calculated as g food/kg body weight/day: 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: No

OPHTHALMOSCOPIC EXAMINATION: No

HAEMATOLOGY: Yes (13-week and 2-year studies only)
- Time schedule for collection of blood: end of the study (13-week studies) or at 15 months (2-year studies)
- Anaesthetic used for blood collection: No
- Animals fasted: No
- How many animals: all (13-week studies) or 5 animals/sex/group (2-year studies)
- Parameters examined: hematocrit, hemoglobin, erythrocytes, mean erythrocyte volume, mean erythrocyte hemoglobin concentration, reticulocytes, total leukocytes and differential, and nucleated erythrocytes

CLINICAL CHEMISTRY: No

URINALYSIS: No

NEUROBEHAVIOURAL EXAMINATION: No

OTHER:

- TISSUE BURDEN ANALYSES: Lungs (rats and mice), kidneys (rats only in 16-day and 13-week studies, or mice only in 2-year studies)

- SPERM MORPHOLOGY AND VAGINAL CYTOLOGY (13-week studies only):
At the end of the studies sperm samples were collected from all male animals in the 0, 0.5, 1 and 2 mglm3 exposure groups for sperm morphology evaluations. The parameters evaluated were sperm density, morphology, and motility. The right epididymis, right caudae, and right testis were weighed. Vaginal samples were collected for up to 7 consecutive days prior to the end of the studies from all female animals in the 0, 0.5, 1, and 2 mglm3 groups for vaginal cytology evaluations. The parameters evaluated were relative frequency of estrous stages and estrous cycle length.

Sacrifice and pathology:

GROSS PATHOLOGY: Yes
Necropsy performed on all animals. Organs weighed (at the 7- and 15-month interim evaluations in the 2-year studies) were brain, heart, right kidney, liver, lung, right testis (rats at 7 months only in the 2-year studies), and thymus.

HISTOPATHOLOGY: Yes

- 16-day studies: Complete histopathology was performed on 0, 30, and 60 mglm3 rats and on 0, 3.5, and 7 mglm3 mice. In addition to gross lesions and tissue masses, the tissues examined included: adrenal gland, bone, brain, clitoral gland (rats only), epididymis or oviduct, esophagus, gallbladder (mice only), heart, large intestine (cecum, colon, rectum), small intestine (duodenum, jejunum, ileum), kidneys, larynx, liver, lung, lymph nodes (bronchial, mandibular, mediastinal, mesenteric), mammary gland, nose, ovary, pancreas, pancreatic islets (rats only), parathyroid gland, pituitary gland, preputial gland (rats only), prostate, salivary gland, seminal vesicle, skin, spleen, stomach (forestomach and glandular), testis, thymus, thyroid gland, trachea, urinary bladder, and uterus. In addition, the following target organs were examined from 3.5, 7, and 15 mglm3 rats: liver, lung, respiratory tract lymph nodes (bronchial and mediastinal), nose, spleen (15 mglm3 only), testis, and thymus.

- 13-week studies: Complete histopathology was performed on 0 and 2 mglm3 rats and mice. In addition to gross lesions and tissue masses, the tissues examined included: adrenal gland, bone, brain, clitoral gland (rats only), epididymis or oviduct, esophagus, gallbladder (mice only), heart, large intestine (cecum, colon, rectum), small intestine (duodenum, jejunum, ileum), kidneys, larynx, liver, lung, lymph nodes (bronchial, mandibular, mediastinal, mesenteric), mammary gland, nose, ovary, pancreas, pancreatic islets (rats only), parathyroid gland, pituitary gland, preputial gland (rats only), prostate, salivary gland, seminal vesicle, skin, spleen, stomach (forestomach and glandular), testis, thymus, thyroid gland, trachea, urinary bladder, and uterus. In addition, the following organs were examined from selected exposure groups of rats and mice: lung, nose, and respiratory tract lymph nodes (bronchial and mediastinal).

- 2-year studies: Complete histopathology was performed on all rats and mice. In addition to gross lesions and tissue masses, the tissues examined included: adrenal gland, bone, brain, clitoral gland, epididymis or oviduct, esophagus, gallbladder (mice only), heart, large intestine (cecum, colon, rectum), small intestine (duodenum, jejunum, ileum), kidneys, larynx, liver, lung, lymph nodes (bronchial, mandibular, mediastinal, mesenteric), mammary gland, nose, ovary, pancreas, pancreatic islets (rats only), parathyroid gland, pituitary gland, preputial gland (except 0 mg\m3 male mice), prostate, salivary gland, seminal vesicle, skin, spleen, stomach (forestomach and glandular), testis, thymus, thyroid gland, trachea, urinary bladder, and uterus.

Other examinations:

No other examinations

Statistics:

Included in the 2-year studies only, for survival analyses, calculation of incidences, analysis of neoplasm incidences, analysis of non-neoplastic lesion incidences, and analysis of continuous variables

Results and discussion

Results of examinations

Clinical signs:

effects observed, treatment-related

Description (incidence and severity):

mortality in 16-day and 13-week studies only; clinical signs in 16-day studies only

Mortality:

mortality observed, treatment-related

Description (incidence):

mortality in 16-day and 13-week studies only; clinical signs in 16-day studies only

Body weight and weight changes:

effects observed, treatment-related

Description (incidence and severity):

in 16-day and 2-year studies only

Food consumption and compound intake (if feeding study):

not examined

Food efficiency:

not examined

Water consumption and compound intake (if drinking water study):

not examined

Ophthalmological findings:

not examined

Haematological findings:

effects observed, treatment-related

Description (incidence and severity):

in 13-week studies only

Clinical biochemistry findings:

not examined

Urinalysis findings:

not examined

Behaviour (functional findings):

not examined

Organ weight findings including organ / body weight ratios:

effects observed, treatment-related

Gross pathological findings:

no effects observed

Histopathological findings: non-neoplastic:

effects observed, treatment-related

Histopathological findings: neoplastic:

no effects observed

Details on results:

16-DAY STUDY IN RATS:

In the core study, two 60 mg/m3 males, one 30 mg/m3 female, and all 60 mg/m3 females died before the end of the study. Final mean body weights of all exposed groups of males and females were significantly lower than those of the controls, as were mean body weight gains of male rats. Clinical findings included increased rates of respiration and reduced activity levels inr ats in all exposure groups, except those exposed to 3.5 mg/m3. Absolute lung weights of 60 mg/m3 males and of all exposed groups of females were significantly greater than those of the controls, as were the relative lung weights of all exposed groups of males and females. Inflammation (including degeneration and necrosis of the bronchiolar epithelium) occurred in the lungs of all exposed groups of males and females. Atrophy of the olfactory epithelium occurred in the nasal passages of all exposed groups of males (except 60 mg/m3) and in 15, 30, and 60 mg/m3 females. Lymphoid hyperplasia in the bronchiolar mediastinal lymph nodes was observed in 30 mg/m3 males and in 60 mg/m3 males and females. The concentration of nickel in the lungs of all exposed groups of males and females was greater than in control animals.

16-DAY STUDY IN MICE:

All core study mice exposed to 7 mg/m3 or greater died before the end of the study; all control and 3.5 mg/m3 mice survived to the end of the study. Final mean body weights and weight gains of 7, 15, 30, and 60 mg/m3 males and females were significantly less than those of the controls, and clinical findings in these groups included emaciation, lethargy, and rapid respiration rates. Absolute and relative lung weights of male and female mice exposed to 7 mg/m3 or greater were significantly greater than those of the controls. Only tissues from mice exposed to 0, 3.5, or 7 mg/m3 were examined histopathologically. Inflammation occurred in the lungs of 3.5 and 7 mg/m3 males and females; necrosis of the alveolar and bronchiolar epithelium was a component of the inflammation in 7 mg/m3 males and females. In addition, atrophy of the olfactory epithelium of the nasal passages was observed in 3.5 mg/m3 males and females. Nickel concentrations in the lungs of mice exposed to 3.5 mg/m3 were greater than those in the controls.

13-WEEK STUDY IN RATS:

In the core study, one 2 mg/m3 male rat died before the end of the study; all other males and all females survived until the end of the study. Final mean body weights and body weight gains of all exposed groups were similar to those of the controls. There were no significant clinical findings noted during the study. Exposure-related increases in neutrophil and lymphocyte counts occurred and were most pronounced in female rats. With the exception of 0.12 mg/m3 rats, absolute and relative lung weights of all exposed groups were generally significantly greater than those of the controls. Exposure-related increases in the incidence and severity of inflammatory lesions (alveolar macrophages, chronic inflammation, and interstitial infiltration) occurred in the lungs of all exposed groups of males and females. Lymphoid hyperplasia of the bronchial and/or mediastinal lymph nodes occurred in males exposed to 0.5 mg/m3 or greater. Atrophy of the olfactory epithelium occurred in males and females exposed to 0.5, 1, and 2 mg/m3 and in 0.25 mg/m3 females. The concentration of nickel in the lungs of 0.5 and 2 mg/m3 rats was greater than that in the lungs of control animals at 4, 9, and 13 weeks for males and at 13 weeks for females.

13-WEEK STUDY IN MICE:

In the core study, four control males, three control females, and one 0.12 mg/m3 male died before the end of the study; the deaths were not considered to be chemical related, and all other mice survived to the end of the study. The final mean body weights and body weight gains of all exposed groups were similar to those of the controls. There were no chemical-related clinical findings. Hematology changes similar to those reported in female rats occurred in female mice, but the mice were minimally affected. The absolute and relative lung weights of 1 mg/m3 males and 2 mg/m3 males and females were significantly greater than those of the controls. Increased numbers of alveolar macrophages occurred in all males and females exposed to 0.5 mg/m3 or greater. Chronic active inflammation and fibrosis occurred in 1 and 2 mg/m3 males and females. Lymphoid hyperplasia of the bronchial lymph node and atrophy of the olfactory epithelium in the nasal passages were observed in 2 mg/m3 males and females. Nickel concentration in the lung of 2 mg/m3 females was significantly greater than in control animals.

2-YEAR STUDY IN RATS:

The survival rates of all exposed groups of males and females were similar to those of the controls. Mean weights of 0.5 mg/m3 female rats were slightly lower (6% to 9%) than those of the controls throughout the second year of the study; final mean body weights of all exposed groups of males and 0.12 and 0.25 mg/m3 females were similar to those of the controls. There were no clinical findings or hematology differences that were considered to be related to nickel sulfate hexahydrate administration.
No exposure-related neoplasms occurred in male or female rats exposed by inhalation to nickel sulfate hexahydrate for 2 years. Increased incidences of inflammatory lung lesions were generally observed in all exposed groups of male and female rats at the end of the study. The incidences of chronic active inflammation, macrophage hyperplasia, alveolar proteinosis, and fibrosis were markedly increased in male and female rats exposed to 0.25 or 0.5 mg/m3. Increased incidences of lymphoid hyperplasia in the bronchial lymph nodes occurred in 0.5 mg/m3 male and female rats at the end of the 2-year study. The incidences of atrophy of the olfactory epithelium in 0.5 mg/m3 males and females were significantly greater than those in controls at the end of the study.
Lung nickel burdens in exposed male and female rats were greater than those in the controls at the 7- and 15-month interim evaluations, and lung nickel burdens values increased with increasing exposure concentration.

2-YEAR STUDY IN MICE:

The survival rates of all exposed groups of males and females were similar to those of the controls. The mean body weights of 1 mg/m3 males and of all exposed groups of females were lower than those of the controls during the second year of the study. There were no clinical findings or hematology differences considered to be related to chemical exposure.
Inflammatory lesions of the lung generally occurred in all exposed groups of male and female mice at the of the 2-year study. These lesions included macrophage hyperplasia, chronic active inflammation, bronchialization (alveolar epithelial hyperplasia), alveolar proteinosis, and infiltrating cells in the interstitium. Incidences of macrophage hyperplasia and/or lymphoid hyperplasia occurred in the bronchial lymph nodes of most of the 1 mg/m3 males and females and in some 0.5 mg/m3 females at the end of the 2-year study. Atrophy of the olfactory epithelium was observed in 0.5 and 1 mg/m3 males and in all exposed groups of females at the end of the 2-year study.
At the 7- and 15-month interim evaluations, lung nickel burden parameters measured in control and exposed groups were below the limit of detection. Absolute lung weights of 0.5 and 1 mg/m3 lung burden study females were significantly greater than those of the controls at 15 months.

Effect levels

1

Dose descriptor:

NOAEC

Remarks:

in 16-day studies in rats and mice, 13-week study in rats, and 2-year studies in rats and mice

Basis for effect level:

other: Organ weight and microscopic changes in lungs and respiratory tract

Remarks on result:

not determinable

Remarks:

no NOAEC identified

2

Dose descriptor:

NOAEC

Remarks:

in 13-week study in mice

Effect level:

0.25 mg/m³ air

Based on:

test mat.

Remarks:

equivalent to 0.06 mg Ni/m3

Sex:

male/female

Basis for effect level:

other: Organ weight and microscopic changes in lungs and respiratory tract

3

Dose descriptor:

LOAEC

Remarks:

in 2-year study in rats

Effect level:

0.25 mg/m³ air

Based on:

test mat.

Remarks:

equivalent to 0.06 mg Ni/m3

Sex:

male/female

Basis for effect level:

other: Increased incidence of lung chronic active inflammation, macrophage hyperplasia, alveolar proteinosis, and fibrosis

Target system / organ toxicity

Critical effects observed:

not specified

Any other information on results incl. tables

The detailed incidences and severities of neoplasms and nonneoplastic lesions in the lungs or rats and mice in the 2 -year studies are provided in Tables 1 and 2 below, respectively.

Table 1: Incidence of lung lesions in rats in the 2-year inhalation study

	Nickel sulfate hexahydrate concentration (mg/m3)
	0		0.12		0.25		0.50
	Males	Females	Males	Females	Males	Females	Males	Females
7-month interim evaluation
Lung:^a	5	5	5	5	5	5	5	5
Inflammation, chronic active^b	0	0	4* (1.0)^c	2 (1.0)	4* (1.3)	4* (1.3)	5** (1.8)	5** (2.0)
Macrophage hyperplasia	0	0	1 (1.0)	2 (1.0)	5** (1.0)	4* (1.0)	5** (2.0)	5** (2.0)
15-month interim evaluation
Lung:	5	5	5	5	5	5	5	5
Inflammation, chronic active	0	2 (1.0)	1 (1.0)	0	1 (1.0)	4 (1.0)	5** (1.0)	5 (1.2)
Macrophage hyperplasia	0	1 (1.0)	0	1 (1.0)	2 (1.0)	3 (1.3)	5** (2.0)	5* (1.8)
Alveolar proteinosis	0	0	0	0	1 (1.0)	3 (1.3)	4* (1.8)	5** (1.4)
Fibrosis	0	0	0	0	0	1 (1.0)	2 (1.0)	3 (1.3)
Hyperplasia, focal	0	1	0	0	1	0	0	1
2-year evaluation
Lung:	54	52	53	53	53	53	53	54
Inflammation, chronic active	14 (1.1)	14 (1.2)	11 (1.2)	13 (1.2)	42** (1.9)	49** (2.1)	46** (2.2)	52** (2.3)
Macrophage hyperplasia	7 (1.3)	9 (1.6)	9 (1.2)	10 (1.1)	35** (1.6)	32** (1.5)	48** (2.2)	45** (1.8)
Alveolar proteinosis	0	1 (1.0)	0	0	12** (1.4)	22** (1.5)	41** (1.9)	49** (2.6)
Fibrosis	3 (1.0)	8 (1.4)	6 (1.2)	7 (1.3)	35** (1.7)	45** (1.7)	43** (1.8)	49** (1.9)
Hyperplasia, focal	3	5	2	3	3	7	2	10
Squamous metaplasia	0	0	0	0	0	0	0	4
Alveolar/bronchiolar adenoma or carcinoma	1	0	0	0	1	0	3	1
* p<0.05              ** p<0.01 ^a  Number of animals with lung examined microscopically ^b  Number of animals with lesion ^c  Average severity grade of lesion in affected animals (1: minimal, 2: mild, 3: moderate, 4:marked)

 

Table 2: Incidence of lung lesions in mice in the 2-year inhalation study

	Nickel sulfate hexahydrate concentration (mg/m3)
	0		0.25		0.50		1.0
	Males	Females	Males	Females	Males	Females	Males	Females
7-month interim evaluation
Lung:^a	5	5	5	5	5	5	5	5
Inflammation, chronic active^b	0	0	0	0	0	0	0	2 (1.0)^c
Macrophage hyperplasia	0	0	0	0	1 (1.0)	1 (1.0)	5** (1.0)	5** (1.0)
Interstitial infiltration	0	0	0	0	0	0	1 (1.0)	1 (2.0)
15-month interim evaluation
Lung:	5	5	5	5	5	5	5	5
Inflammation, chronic active	0	0	0	0	0	0	4* (1.3)	5** (1.2)
Bronchialization	0	0	1 (1.0)	0	0	1 (1.0)	5** (1.0)	5** (1.0)
Macrophage hyperplasia	0	0	1 (1.0)	1 (1.0)	4* (1.0)	2 (1.0)	5** (1.0)	5** (1.2)
Interstitial infiltration	0	1 (1.0)	0	0	0	0	5** (1.0)	5* (1.2)
Alveolar proteinosis	0	0	0	0	0	0	3 (1.0)	5** (1.2)
2-year evaluation
Lung:	61	61	61	60	62	60	61	60
Inflammation, chronic active	1 (2.0)	1 (1.0)	2 (1.0)	7* (1.3)	8* (1.5)	14** (1.1)	29** (1.9)	40** (1.6)
Bronchialization	1 (1.0)	0	4 (1.5)	9** (1.0)	19** (1.1)	32** (1.0)	39** (1.1)	45** (1.1)
Macrophage hyperplasia	6 (2.8)	7 (1.6)	9 (1.4)	24** (1.3)	35** (1.5)	53** (1.5)	59** (2.1)	59** (2.4)
Interstitial infiltration	1 (3.0)	0	0	4 (2.0)	3 (1.3)	16** (1.1)	17** (1.2)	39** (1.4)
Alveolar proteinosis	0	0	0	0	0	11** (1.4)	42** (2.0)	45** (1.9)
Alveolar epithelial hyperplasia, focal	0	0	0	1 (2.0)	0	1 (2.0)	0	0
Alveolar/bronchiolar adenoma	5	3	5	3	3	2	5	0
Alveolar/bronchiolar carcinoma	9	4	13	3	4	9	3	2
* p<0.05              ** p<0.01 ^a  Number of animals with lung examined microscopically ^b  Number of animals with lesion ^c  Average severity grade of lesion in affected animals (1: minimal, 2: mild, 3: moderate, 4:marked)

 

Applicant's summary and conclusion

Conclusions:

Under the conditions of these 2-year inhalation studies, there was no evidence of carcinogenic activity of nickel sulfate hexahydrate in male or female F344/N rats exposed to 0.12, 0.25, or 0.5 mg/m3 (0.03, 0.06, or 0.11 mg nickel/m3). There was no evidence of carcinogenic activity of nickel sulfate hexahydrate in male or female B6C3F, mice exposed to 0.25, 0.5, or 1 mg/m3 (0.06, 0.11, or 0.22 mg nickel/m3). Target organ for non-cancer toxicity was lung, with clear concentration-related chronic active inflammation and fibrosis observations.

Executive summary:

Male and female F344/N rats and B6C3F1 mice were exposed to nickel sulfate hexahydrate (mass mean diameter 1.8 -3.1 µm +/- 1.6 -2.9; greater than 98% pure) by whole-body inhalation exposure for 16 days or 13 weeks (as preliminary studies), or 2 years (6 hours a day, 5 days a week). The concentrations used in the 2 -year studies were 0, 0.12, 0.25, or 0.5 mg/m3 (equivalent to 0, 0.03, 0.06, or 0.11 mg nickel/m3) and 0, 0.25, 0.5, or 1 mg/m3 (equivalent to 0, 0.06, 0.11, or 0.22 mg nickel/m3) , in rats and mice, respectively.

Mortality occurred in both species in the 16 -day and 13 -week studies. The survival rate of exposed animals was similar to that of controls in the 2 -year studies. Clinical signs (increased respiratory rate, reduced activity, emaciation, lethargy) were observed in both species in the 16 -day studies only. Significant decreases in body weight and/or body weight gain were noted following 16 days or 2 years of exposure in both species. Relevant changes in hematology occurred in both species in the 13 -week studies only, and consisted of increased neutrophil and lymphocyte counts, more markedly for females. Increased absolute and/or relative lung weights were observed in both species whatever the duration of exposure. There were no relevant effects on gross pathology. No neoplastic microscopic abnormalities were observed. Histopathological examination revealed microscopic inflammatory changes in the lungs of both species whatever the duration of exposure.

Exposure of rats to nickel sulfate hexahydrate by inhalation for 2 years resulted in increased incidences of chronic active inflammation, macrophage hyperplasia, alveolar proteinosis, and fibrosis of the lung; lymphoid hyperplasia of the bronchial lymph node; and atrophy of the olfactory epithelium. Exposure of mice to nickel sulfate hexahydrate by inhalation for 2 years resulted in increased incidences of chronic active inflammation, bronchialization (alveolar epithelial hyperplasia), macrophage hyperplasia, interstitial infiltration, and alveolar proteinosis of the lung; lymphoid and macrophage hyperplasia of the bronchial lymph node; and atrophy of the olfactory epithelium. There was no evidence of carcinogenic activity in either rodent species.

In these repeat-dose inhalation toxicity studies in rats and mice, the lung was clearly a target organ of nickel sulfate hexahydrate toxicity. Based on organ weight and microscopic changes in lungs and respiratory tract, no NOAEC can be identified in any study, except in the 13 -week study in mice where 0.25 mg/m3 (equivalent to 0.06 mg nickel/m3) was a NOAEC. In particular following chronic exposure in rats for 2 years (6 hours a day, 5 days a week), the concentration of 0.25 mg/m3 (equivalent to 0.06 mg nickel/m3) can be considered as a LOAEC based on the increased incidence of chronic active inflammation, macrophage hyperplasia, alveolar proteinosis, and fibrosis in male and female rats.