Registration Dossier

Administrative data

Description of key information

Oral Route:

No repeated oral dose toxicity data of sufficient quality are available for fused tungsten carbide (target substance). However, reproductive toxicity data are available for sodium tungstate (source substance), which will be used for read-across. Due to lower water solubility and lower toxicity for the target substance compared to the source substance, the resulting read-across from the source substance to the target substance is appropriate as a conservative estimate of potential toxicity for this endpoint. In addition, read-across is appropriate because the classification and labelling is more protective for the source substance than the target substance, the PBT/vPvB profile is the same, and the dose descriptors are, or are expected to be, lower for the source substance. For more details, refer to the attached description of the read-across approach.

A repeated dose study on sodium tungstate was sponsored conducted the United States Army Center for Health Promotion and Preventive Medicine and published by McCain et al (2015). The 90 -day oral toxicity study was conducted in rats according to the procedure described in the Environmental Protection Agency (EPA) Health Effects Testing Guidelines (40 CFR, Part 798.2650) in compliance with Good Laboratory Practice. Briefly, this study of the subchronic toxicity of sodium tungstate dihydrate aqueous solution in male and female Sprague-Dawley rats was evaluated by daily oral gavage of 0, 10, 75, 125, or 200 mg/kg bw/d for 90 days. Measured parameters included food consumption, body weight measurements, hematology, clinical chemistry, and histopathological changes. There was a significant decrease in food consumption and body weight gain in males at 200 mg/kg bw/d from days 77 to 90; however, there was no effect in food consumption and body weights in females. There were no changes in the hematological and clinical parameters studied. Histopathological changes were seen in kidney of male and female and epididymis of male rats. The histopathological changes observed in the kidneys of male and female rats dosed at 125 or 200 mg/k/d consisting of mild to severe cortical tubule basophilia in 2 high-dose groups. Histological changes in epididymides included intraluminal hypospermia with cell debris in the 200 mg/kg bw/day dosed male rats. Histopathological changes were observed in the glandular stomach including inflammation and metaplasia in the high-dose groups (125 or 200 mg/kg bw/day) of both sexes of rats. Based on histopathology effects seen in the kidneys, the lowest observable adverse effect level was 125 mg/kg bw/d and the no observable adverse effect level was 75 mg/kg bw/d in both sexes of rats for oral subchronic toxicity. The USEPA’s Benchmark Dose Software (BMDS, Version 1.4.1) was used to model the data to derive a BMDL10. The lowest (most precautionary) BMDL10 from the renal toxicity endpoint in the 90-day oral toxicity study was 102 mg/kg bw/d.

Repeated Dose Toxicity - Inhalation Route:

No inhalation repeated dose toxicity data of sufficient quality are available for fused tungsten carbide (target substance). However, inhalation repeated dose toxicity data are available for tungsten oxide (source substance), which will be used for read-across. Due to similar water solubility and lower toxicity for the target substance compared to the source substance, the resulting read-across from the source substance to the target substance is appropriate. In addition, read-across is appropriate because the classification and labelling is similar for the source substance than the target substance, the PBT/vPvB profile is the same, and the dose descriptors are, or are expected to be, lower for the source substance. For more details, refer to the read-across category approach included in the Category section of this IUCLID submission and/or as an Annex in the CSR.

A 28-day inhalation toxicity study conducted according to OECD 412 is available on tungsten oxide (also known as tungsten blue oxide or TBO), which is considered the key repeated dose study. In this study, 5 rats/sex/dose were given TBO nose-only for 6 hours per day, 7 days/week, for 28 days (with a 14-day recovery period) at doses of 0 (control), 0.08, 0.325, and 0.65 mg/L air. The NOAEC was deemed to be > 0.65 mg/L air (650 mg/m3), as no significant effects were reported.

Key value for chemical safety assessment

Toxic effect type:
dose-dependent

Repeated dose toxicity: via oral route - systemic effects

Link to relevant study records
Reference
Endpoint:
sub-chronic toxicity: oral
Type of information:
read-across based on grouping of substances (category approach)
Adequacy of study:
key study
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Remarks:
Well documented, scientifically sound study that was conducted according to EPA OTS 798.2650 (90-Day Oral Toxicity in Rodents)
Justification for type of information:
1. HYPOTHESIS FOR THE CATEGORY APPROACH: The hypothesis is that properties are likely to be similar or follow a similar pattern because of the presence of a common metal ion, in this case tungstate.
2. SOURCE AND TARGET CHEMICAL(S) (INCLUDING INFORMATION ON PURITY AND IMPURITIES):
Source: Sodium tungstate
Target: Fused tungsten carbide
3. CATEGORY APPROACH JUSTIFICATION: See Annex 3 in CSR
4. DATA MATRIX: See Annex 3 in CSR
Reason / purpose for cross-reference:
assessment report
Reason / purpose for cross-reference:
read-across source
Qualifier:
according to guideline
Guideline:
EPA OTS 798.2650 (90-Day Oral Toxicity in Rodents)
Deviations:
no
GLP compliance:
yes
Limit test:
no
Specific details on test material used for the study:
Sodium tungstate dihydrate (Tungstic acid sodium salt dihydrate, Na2WO42H2O, CAS # 10213-10-2, Batch # 12330JO) 99% pure
Species:
rat
Strain:
other: SD
Details on species / strain selection:
5-week-old SD rats were purchased from Charles River laboratories, Raleigh, North Carolina
Sex:
male/female
Details on test animals or test system and environmental conditions:
Animals were held for 1 week in quarantine prior to initiation of treatments. At the start of testing, rats weighed between 199 and 230 g. Test animals were identified by individual cage cards and microchip implants and were individually housed in
polycarbonate cages. Bedding was placed in the bottom of each cage and replaced twice weekly. Drinking quality water and a certified laboratory diet were available ad libitum. Animal rooms were maintained at 64 F to 79 F, with relative humidity of 30% to 70% and a 12-hour light–dark cycle.
Route of administration:
oral: drinking water
Details on route of administration:
Sodium tungstate dihydrate was solubilized with DI water to produce 4 dosing solutions
Vehicle:
water
Details on oral exposure:
The dose levels were selected on the basis of our previous studies where the highest dose used was 200 mg/kg/ d in a subchronic toxicity study. Sodium tungstate dihydrate was solubilized with DI water to produce 4 dosing solutions of
200, 125, 75, and 10 mg Na2WO4/mL.
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
Tungstate concentrations of the dosing solutions were verified by the Aberdeen Test Center and found to be consistent for purity and stability during the study period
Duration of treatment / exposure:
A 90-day oral toxicity study
Frequency of treatment:
Test chemical solutions were administered daily (7 days per week) for 90 days. A 16 GA 2-in stainless steel gavage needle
Dose / conc.:
10 mg/kg bw/day (actual dose received)
Dose / conc.:
75 mg/kg bw/day (actual dose received)
Dose / conc.:
125 mg/kg bw/day (actual dose received)
Dose / conc.:
200 mg/kg bw/day (actual dose received)
No. of animals per sex per dose:
Following 1-week quarantine/acclimatization period, 50 male and 50 female SD rats were randomly distributed
Control animals:
yes, concurrent vehicle
Details on study design:
Sodium tungstate dihydrate was solubilized with DI water to produce 4 dosing solutions of 200, 125, 75, and 10 mg Na2WO4/mL.
Observations and examinations performed and frequency:
A clinical examination was made for each animal prior to initiation of treatment and once weekly during treatment. Observations included but were not limited to changes in skin and fur, eyes, mucous membranes, occurrence of secretions and excretions, and autonomic activity (eg, lacrimation, piloerection, pupil size, and unusual respiratory pattern). Changes in gait, posture, and response to handling as well as the presence of clonic or tonic movements, stereotypes (eg, excessive grooming, repetitive circling), or bizarre behavior (eg, selfmutilation, walking backward) were recorded according to testing guidelines. Body and feeder weights were recorded on days .3, .1, 0 (first day of dosing), 7, and weekly thereafter. Doses were adjusted weekly to reflect the change in individual body weights. Animals were observed daily for any toxic signs.
Sacrifice and pathology:
Blood was collected. Each rat was then submitted for complete necropsy. The brain, heart, liver, kidneys, spleen, adrenals, thymus, epididymis/uterus, and testes/ovaries were removed and weighed for absolute organ weights. The tissues harvested for histopathological evaluation included the brain, pituitary, thyroid with parathyroid gland, thymus, lungs, trachea, heart, bone marrow, salivary gland, liver, spleen, kidney, adrenal gland, pancreas, testis, ovaries, uterus, aorta, esophagus, stomach, duodenum, jejunum, ileum, caecum, colon, urinary bladder, salivary lymph node peripheral nerve (siatic), thigh musculature (vastus lateralis), eye, spinal cord (3 levels), and exorbital lachrymal gland.

Hematology parameters included white blood cell count, neutrophils, lymphocytes, monocytes, eosinophils, basophils, red blood cell count, hemoglobin, hematocrit, mean cell volume, mean cell hemoglobin, mean cell hemoglobin concentration, red blood cell distribution width, platelets, and mean platelet volume.

Clinical chemistry parameters measured included The clinical chemistry analytes ialkaline phosphatase, alanine aminotransferase, aspartate aminotransferase, blood urea nitrogen, calcium, cholesterol, creatinine kinase, creatinine, glucose (GLU; nonfasting), lactate dehydrogenase, total bilirubin, total protein, triglycerides, Na, K, and Cl

Urinalysis included appearance, pH, specific gravity, GLU, bilirubin, urobilinogen, ketone, blood, protein, nitrite, and leukocytes.
Other examinations:
Ophthalmic examinations were performed on all control and treated animals prior to the scheduled start of the study and within a week of the scheduled 90-day necropsies. Urinalysis was also performed on 8 of 10 animals from all dose
groups (including negative control) within 2 weeks of the final (90-day) necropsies
Statistics:
Food consumption, body weights, and absolute organ weights were compared among dosage groups and controls using ANOVA. When significance was observed, the data were further analyzed using a Dunnett test to compare the doses to the control group. Statistical significance was defined at the P <05 level. Clinical chemistry, hematology, and urinalysis data were analyzed with ANOVA and Bonferroni post hoc test to compare dosage groups to the control group.
Clinical signs:
no effects observed
Description (incidence and severity):
- No evidence of overt toxicity and no treatment-related clinical signs were seen in any dose levels.
- Results showed that rats dosed with sodium tungstate in water for 90 consecutive days had no abnormal clinical signs at any of the dose levels
Mortality:
mortality observed, non-treatment-related
Description (incidence):
No unscheduled deaths (a single male rat in 200 mg/kg/d dose group was moribund and was euthanized on day 79; a few tissues from this rat were submitted for histopathological examination and death was determined to be not compound related
Body weight and weight changes:
effects observed, treatment-related
Description (incidence and severity):
There were significant decreases in food consumption in male rats at 200 mg/kg/d from weeks of 4 to 13, while there was no change in food consumption in female rats during the 90-day study
Food consumption and compound intake (if feeding study):
effects observed, treatment-related
Description (incidence and severity):
There were significant decreases in food consumption in male rats at 200 mg/kg/d from weeks of 4 to 13, while there was no change in food consumption in female rats during the 90-day study changes in female rats in any dose groups throughout study
period when compared to control rats.
Food efficiency:
not specified
Water consumption and compound intake (if drinking water study):
not specified
Ophthalmological findings:
no effects observed
Description (incidence and severity):
Ophthalmic examinations prior to study initiation and within
a week of the scheduled necropsies revealed no abnormalities
Haematological findings:
no effects observed
Description (incidence and severity):
Male and female rats showed no significant differences in any hematological parameters at any dose levels of sodium tungstate
Clinical biochemistry findings:
no effects observed
Description (incidence and severity):
The results of clinical chemistry parameters studied in rats showed no significant changes in any dose levels of sodium tungstate in rats. The parameters studied showed some changes in levels that were not dose related and insignificant and considered within normal range limits when compared to controls. All other parameters were found to be similar to control rats.

Urinalysis findings:
no effects observed
Description (incidence and severity):
Examination of urine samples taken approximately 1 week prior to necropsy revealed no significant changes in volume, specific gravity, or pH. No distinct dose-related trends were observed in GLU, bilirubin, ketone, blood, protein, urobilinogen,
nitrite, or leukocytes
Behaviour (functional findings):
no effects observed
Immunological findings:
not examined
Organ weight findings including organ / body weight ratios:
effects observed, treatment-related
Description (incidence and severity):
The body weights, absolute heart, liver, and thymus weights were significantly lower in male rats dosed at 200 mg/kg/d compared to control rat, but there were no effects on body weights and organ weights of female rats
Gross pathological findings:
not specified
Neuropathological findings:
not examined
Histopathological findings: non-neoplastic:
effects observed, treatment-related
Description (incidence and severity):
- Histopathology examination revealed effects on the urogenital system of the sodium tungstate-treated rats. Changes included mild to severe basophilia of renal cortical tubules in 1 of 9 and 10 of 10 males and 1 of 10 and 8 of 10 females in 2 high-dose
groups (125 and 200 mg/kg/d), respectively.
- Histopathological analysis of epididymides of rats dosed with sodium tungstate showed considerable effects in the high-dose group, Intraluminal cellular debris with and without hypospermia was noted in the epididymides of 3 of 10 males in the 200 mg/kg/d dose group. The lesion was not observed in the 10, 75, and 125 mg/kg/d dose groups.
- Histologic changes were also noted in the glandular stomach of males and females in high dosage groups. The changes included subacute inflammation consisting primarily of EOSs admixed with fewer mononuclear cells observed throughout the submucosa of 5 of 9, 4 of 10 males, and 8 of 10, 9 of 10 females in, 125 and 200 mg/kg/d dosage groups, respectively. Goblet cell metaplasia was also observed in the mucosa of the glandular stomach 8 of 9, 8 of 10 males and 8 of 10, 10 of 10
females of 125 and 200 mg/kg/d dosage groups, respectively. The gastric histologic findings in the lower dosed group were negative when compared to 2 high-dose groups
Histopathological findings: neoplastic:
no effects observed
Key result
Dose descriptor:
BMDL10
Effect level:
102 mg/kg bw/day (actual dose received)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
histopathology: non-neoplastic
Key result
Dose descriptor:
NOAEL
Effect level:
75 mg/kg bw/day (actual dose received)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
histopathology: non-neoplastic
Key result
Dose descriptor:
LOAEL
Effect level:
125 mg/kg bw/day (actual dose received)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
histopathology: non-neoplastic
Dose descriptor:
LOAEL
Effect level:
175 mg/kg bw/day (actual dose received)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
histopathology: non-neoplastic
Key result
Dose descriptor:
LOAEL
Effect level:
200 mg/kg bw/day (actual dose received)
Based on:
test mat.
Sex:
male
Basis for effect level:
food consumption and compound intake
Key result
Dose descriptor:
LOAEL
Effect level:
200 mg/kg bw/day (actual dose received)
Based on:
test mat.
Sex:
male
Basis for effect level:
body weight and weight gain
Key result
Dose descriptor:
NOAEL
Effect level:
200 mg/kg bw/day (actual dose received)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
clinical signs
haematology
Key result
Dose descriptor:
LOAEL
Effect level:
200 mg/kg bw/day (actual dose received)
Based on:
test mat.
Sex:
male
Basis for effect level:
histopathology: non-neoplastic
Key result
Critical effects observed:
yes
Lowest effective dose / conc.:
125 mg/kg bw/day (actual dose received)
System:
urinary
Organ:
kidney
Treatment related:
yes
Dose response relationship:
yes
Relevant for humans:
yes
Conclusions:
Subchronic toxicity of sodium tungstate was assessed in in male and female Sprague-Dawley rats by daily oral gavage of 0, 10, 75, 125, or 200 mg/kg/d for 90 days. There was a significant decrease in food consumption and body weight gain in males at 200 mg/kg/d from days 77 to 90; however, there was no effect in food consumption and body weights in females. There were no changes in the hematological and clinical parameters studied. Histopathological changes were seen in kidney of male and female and epididymis of male rats. Histopathological changes were observed in the kidneys of male and female rats dosed at 125 or 200 mg/k/d consisting of mild to severe cortical tubule basophilia in 2 high-dose groups. Histological changes in epididymides included intraluminal hypospermia with cell debris in the 200 mg/kg/d dosed male rats. Histopathological changes were observed in the glandular stomach including inflammation and metaplasia in the high-dose groups (125 or 200 mg/kg/d) of both sexes of rats. Based on histopathology effects seen in the kidneys, the lowest observable adverse effect level was 125 mg/kg/d and the no observable adverse effect level was 75 mg/kg/d in both sexes of rats for oral subchronic toxicity.
Executive summary:

No oral repeated dose toxicity data of sufficient quality are available for fused tungsten carbide (target substance). However, oral repeated dose toxicity data are available for sodium tungstate (source substance), which will be used for read-across. Due to lower water solubility and lower toxicity for the target substance compared to the source substance, the resulting read-across from the source substance to the target substance is appropriate as a conservative estimate of potential toxicity for this endpoint. In addition, read across is appropriate because the classification and labelling is more protective for the source substance than the target substance, the PBT/vPvB profile is the same, and the dose descriptors are, or are expected to be, lower for the source substance. For more details, refer to the read-across category approach included in the Category section of this IUCLID submission and/or as an Annex in the CSR.

Endpoint conclusion
Endpoint conclusion:
adverse effect observed
Dose descriptor:
BMDL10
102 mg/kg bw/day
Study duration:
subchronic
Species:
rat
Quality of whole database:
The reliability of this study for the substance tested is a K1
System:
urinary
Organ:
kidney

Repeated dose toxicity: inhalation - systemic effects

Link to relevant study records
Reference
Endpoint:
short-term repeated dose toxicity: inhalation
Type of information:
read-across based on grouping of substances (category approach)
Adequacy of study:
key study
Study period:
2010-08-31 to 2010-01-29
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
guideline study with acceptable restrictions
Remarks:
The reliability of this study for the substance tested is a K1, but in application of read-across to a different substance ECHA’s guidance specifies that the score can be a maximum of K2. Due to higher water solubility and greater in vitro bioaccessibility in synthetic alveolar, lysosomal, and interstitial fluids simulating inhalation exposure for the source substance (TBO) as compared to the target substance (tungsten carbide) and lack of toxicity from acute toxicity studies for the target and source substances, toxicity data on the source substance is expected to represent a worse case, so read-across is appropriate between these substances. In addition, read-across is appropriate for this endpoint because the classification and labelling for human health toxicity endpoints is the same for the source and target substances, the PBT/vPvB profile is the same, and the dose descriptors are, or are expected to be, sufficiently similar or lower for the source substance. For more details, refer to the attached description of the read-across approach.
Justification for type of information:
1. HYPOTHESIS FOR THE CATEGORY APPROACH: The hypothesis is that properties are likely to be similar or follow a similar pattern because of the presence of a common metal ion, in this case tungstate.
2. SOURCE AND TARGET CHEMICAL(S) (INCLUDING INFORMATION ON PURITY AND IMPURITIES):
Source: Tungsten Oxide
Target: Fused Tungsten Carbide
3. CATEGORY APPROACH JUSTIFICATION: See Annex 3 in CSR
4. DATA MATRIX: See Annex 3 in CSR
Reason / purpose for cross-reference:
reference to same study
Reason / purpose for cross-reference:
read-across: supporting information
Qualifier:
according to guideline
Guideline:
OECD Guideline 412 (Subacute Inhalation Toxicity: 28-Day Study)
GLP compliance:
yes
Limit test:
no
Species:
rat
Strain:
Sprague-Dawley
Sex:
male/female
Details on test animals or test system and environmental conditions:
TEST ANIMALS
- Source: Charles River Laboratories, (St. Constant, Canada)
- Age at study initiation: approximately 8 weeks
- Weight at study initiation: One day following receipt, body weight ranges of the first shipment of rats 226 to 279 g (males) and 146 to 173 g (females). One day following receipt, body weight range of the second shipment of male rats was 207 to 234 g.
- Fasting period before study: no food or water was provided during exposures.
- Housing: At the start of food consumption measurements, the rats were individually housed in clear polycarbonate rodent cages (Allentown Caging Equipment Co., Allentown, NJ).
-Diet: Certified Rodent Chow 5002 meal (PMI Nutrition International, Inc., Brentwood, MO) was provided ad libitum, except during inhalation exposures and scheduled fasting periods. Diet analysis reports received from the supplier are maintained with facility records. The diet contained no known contaminants at levels that would be expected to interfere with the test substance or the animals or confound interpretation of the study.
- Water (e.g. ad libitum): Each rodent cage was provided with an automatic watering system (Edstrom Industries, Inc., Waterford, WI) supplying fresh city of Chicago water without additional treatment ad libitum, except during inhalation exposures.
- Acclimation period: The animals were quarantined for 2 weeks; To condition the animals for placement and restraint in the nose-only exposure tubes, and reduce stress during the exposure phase, the animals were acclimated to the restraining tubes during a three-day acclimation period. Animals were restrained for 1/4 (1.5 hours), 1/2 (3 hours), and 3/4 (4.5 hours) of the daily exposure duration (6 hours) on three non-holiday weekdays before the animals were exposed.


ENVIRONMENTAL CONDITIONS
- Temperature (°C): 18.6 to 23.0 degree C
- Humidity (%): 25.1-64.6%
- Air changes (per hr): no data
- Photoperiod (hrs dark / hrs light): automatic 12-hour light/dark cycle was maintained in the exposure and housing chamber laboratories.



IN-LIFE DATES: From: 2010-09-09 To: 2010-10-21
Route of administration:
inhalation: dust
Type of inhalation exposure:
nose only
Vehicle:
other: unchanged (no vehicle)
Remarks on MMAD:
MMAD / GSD: The mean MMADs of the test atmosphere were 2.63, 2.87 and 2.74 um with GSDs of 1.89, 1.94 and 1.92 for Groups 2 through 4, respectively.

Details on inhalation exposure:
GENERATION OF TEST ATMOSPHERE / CHAMBER DESCRIPTION
- Exposure apparatus: The nose-only chamber employed for test substance exposure was contained in an acrylic enclosure to isolate the exposure chamber and protect laboratory personnel. The dilution air to the atmosphere generator was of breathable quality and was filtered with a compressed air filter and a carbon absorber. The exhaust from the exposure chamber was moved through a particulate filter by a ring compressor and exhausted outside the building. Inlet and exhaust flows to and from the chamber were continuously monitored by rotameters.
- Method of holding animals in test chamber: During the inhalation exposures, the rats were restrained in nose-only exposure animal holding tubes (CH Technologies, Westwood, NJ). Animal tube loading and unloading, and tube insertion and removal from the exposure chamber were performed
according to standard procedures designed to minimize stress to study rats. At all times that rats were restrained in holders, they were observed
frequently and when necessary, action was taken to avoid injury, death, or improper exposure. Prior to the start of the exposure, rats were transferred from their housing cages to the nose-only holding tubes. Following confirmation of correct animal number, the animals in the holders were inserted into the ports of the exposure chambers. Following the exposure, the holders were removed. The rats were removed from the holders and returned to their home cages. Chamber port rotation occurred weekly.
- System of generating particulates/aerosols: Test atmospheres in the exposure chambers were generated by aerosolizing the test substance using a compressed air-operated Wright Dust Aerosol Generation System positioned over the chamber. Each inhalation exposure system was equipped with a separate aerosol generation system. The test substance was weighed out and packed into a dust reservoir daily. A constant speed rotating scraper separated a thin film of the test substance at the surface of the cake and delivered it into a dispersing unit, drawn in by aspiration and dispersed by a high velocity air jet. The resulting test atmosphere entered a mixing plenum where it was diluted with breathable quality compressed air to the target concentration prior to introduction to the nose-only inhalation exposure chamber.
- Air flow rate: The total airflow was set to produce an airflow range of approximately 0.5 to 1.0 L/min/exposure port.
- Method of particle size determination: The aerosol particle size distribution was monitored twice per week during the exposure phase of the study by an Aerodynamic Particle Sizer (APS) 3321 with Aerosol Diluter 3302A (both manufactured by TSI Inc., Shoreview, MN). The APS sizes particles in the range from 0.5 to 20 um using a time-of-flight technique that measures aerodynamic diameter in real time.


TEST ATMOSPHERE
- Brief description of analytical method used: The test atmosphere mass concentration was monitored gravimetrically by collecting gravimetric samples on pre-weighed glass fiber filters placed in closed-face filter holders. Samples were collected at a constant flow rate equal to the port flow of the delivery tube, and the total volume of air sampled was measured by a dry gas meter. Test atmosphere samples were collected at least three times during the exposure (generally, once during the first two hours, once during the middle two hours and once during the last two hours). The filter-collected samples were weighed and one filter per group per day (including the control to confirm the absence of test substance in the test atmosphere) was analyzed chemically to confirm the mass of TBO collected; percent recovery (chemical analysis concentration vs. gravimetric concentration) was calculated for each filter analyzed. Chemical analysis was conducted by means of ICP-mass spectrometry. In addition, the test atmosphere aerosol concentration in each chamber was monitored with a real-time aerosol sensor (model # pDR-1000AN, MIE, Inc. Bedford, MA). The sensors were employed only as a real-time indicator of short-term changes in aerosol concentration and were used in guiding laboratory personnel if concentration excursions were encountered.
- Samples taken from breathing zone: yes
Analytical verification of doses or concentrations:
yes
Duration of treatment / exposure:
6 hours/day, 7 days/week for 28 days (14-day recovery period)
Frequency of treatment:
daily
Remarks:
Doses / Concentrations:
0.08, 0.325 and 0.65 mg/L Air
Basis:
other: Target TBO Concentration
Remarks:
Doses / Concentrations:
15.2, 61.8, 123.6 mg/kg/day
Basis:
other: Target Inhaled Concentration (calculated)
Remarks:
Doses / Concentrations:
0.081, 0.331, and 0.652 mg/L
Basis:
other: mean concentrations determined gravimetrically
Remarks:
Doses / Concentrations:
14.8, 60.2, and 118.8 mg/kg/day
Basis:
other: mean inhaled concentrations (calculated)
No. of animals per sex per dose:
5 animals/sex/group
Control animals:
yes, sham-exposed
Details on study design:
- Dose selection rationale:
- Rationale for animal assignment: Rats were assigned to groups using a computerized randomization procedure based on body weights using a "measure random" method that will produce similar group mean values.
- Group 1: Control
- Group 2: Low dose group
- Group 3: Mid dose group
- Group 4: High dose group
- Rationale for selecting satellite groups: Designated Subgroup
- Post-exposure recovery period in satellite groups: 14 days
Positive control:
no
Observations and examinations performed and frequency:
CAGE SIDE OBSERVATIONS: Yes
- Time schedule: The toxicology animals were observed for mortality and moribundity once daily during quarantine, twice daily during the exposure period (once in the morning and once in the afternoon) and once daily during the recovery period.


DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: All animal groups (core and recovery) received a thorough clinical examination daily during the study. Clinical observations during the exposure period were recorded before the exposure and within one hour after exposure termination following removal from the exposure chamber and holding tube. All clinical signs of altered behavior, changes in coat condition, unusual discharge of body fluid, abnormal respirations, lesions or other relevant findings were recorded. Clinical observations were recorded using ToxData© System, version 2.1.E.5 (Pathology Data Systems, Basel, Switzerland).


BODY WEIGHT: Yes
- Time schedule for examinations: Body weights of all animals were determined one day after receipt and on the day of randomization to facilitate test subject selection. Body weights were measured for core and recovery animals on study days 1, 8, 15, 22, 28 (body weights for recovery animals were measured on study Day 29); fasted body weight measured on Day 29 (core animals only). During the recovery period, body weights were measured on study Days 36, 42 and a fasted body weight on study Day 43. Body weight measurements were collected using ToxData© System, version 2.1.E.5.


FOOD CONSUMPTION:
- Food consumption for each animal was determined and mean daily diet consumption calculated as g food/kg body weight/day: Yes; Food consumption for the core and recovery animals was measured five days prior to the first exposure (included in the study data but not reported), and daily on study Days 1, 8, 15, 22 and 29. During the recovery period, food consumption was measured on study Days 36 and 42. Food consumption was collected using ToxData© System, version 2.1.E.5.


HAEMATOLOGY: Yes
- Time schedule for collection of blood: Performed on all core animals on study Day 29 and all recovery animals on study Day 43.
- Anaesthetic used for blood collection: Yes; 70% CO2/30% O2
- Animals fasted: Yes
- How many animals: All animals
- Parameters examined: Red blood cell count and morphology, hematocrit, hemoglobin concentration, mean corpuscular volume, mean corpuscular hemoglobin, mean corpuscular hemoglobin concentration, total and differential leukocyte count (absolute and relative), reticulocyte count (absolute
and relative) and platelet count. The following coagulation parameters were evaluated: fibrinogen, prothrombin time and activated partial thromboplastin time. The data was evaluated using a Diagnostica Stago STA Compact CT Coagulation Analyzer (DIAGNOSTICA STAGO, Inc., Parsippany, NJ), the Coagulation data were transferred from the Coagulation Analyzer to ToxData System, version 2.1.E.5.


CLINICAL CHEMISTRY: Yes
- Time schedule for collection of blood: Performed on all core animals on study Day 29 and all recovery animals on study Day 43.
- Animals fasted: Yes
- How many animals: All animals
- Parameter examined: alanine aminotransferase, albumin, alkaline phosphatase, aspartate aminotransferase, total bilirubin, blood urea nitrogen, calcium, chloride, cholesterol, creatinine, gamma-glutamyl transpeptidase, glucose, lactate dehydrogenase, phosphorus, potassium, sodium, total protein and triglycerides.


URINALYSIS: Yes
- Time schedule for collection of urine: Performed on all core animals on study Day 29 and all recovery animals on study Day 43.
- Metabolism cages used for collection of urine: No data
- Animals fasted: No data
- Parameters examined: pH, protein, glucose, ketones, occult blood, bilirubin, urobilinogen, nitrite and leukocytes. Refractive index was measured with a refractometer, and a species-specific urine solids table was used to convert the refractive index to specific gravity. Urine was evaluated macroscopically for color, clarity and volume, and the sediment was analyzed microscopically.


OTHER:
ORGAN WEIGHTS: At the terminal and recovery necropsies, the adrenal glands, brain, epididymides, paired kidneys, liver, lungs, spleen, uterus, thymus and paired gonads (testes or ovaries) were removed, trimmed, and weighed. Organ weight/body weight ratios (relative organ weights) were calculated using the fasted body weights obtained prior to necropsy.

Sacrifice and pathology:
GROSS PATHOLOGY: Yes; complete necropsies were conducted on all animals at terminal and recovery sacrifice. The necropsy included examination of the external surface and all orifices; all body cavities including the cranial cavity; and collection and fixation of the following tissues and organs: adrenal glands, brain, epididymides, heart (with aorta), kidneys, liver (two sections including left lateral and median lobes), lungs (with mainstem bronchi), ovaries, pancreas, seminal vesicles, spleen, sternum (bone marrow), testes, thymus and uterus. Gross lesions were also collected and preserved from all animals. In addition, the section of the tail bearing the animal identification number was also collected and preserved from all animals, and two bone marrow smears were collected from the femur of each animal.

HISTOPATHOLOGY: Yes; The adrenal glands, kidneys, liver (two sections including left lateral and median lobes), lungs (with mainstem bronchi), spleen and any gross lesions for the control and high dose groups were processed for microscopic evaluation for all core and recovery animals at necropsy (except as noted in Protocol Deviation No. 1). Tissues from other dose group animals were saved for future evaluations. Lungs (with mainstem bronchi) for the low and mid dose groups were processed for microscopic evaluation for all core toxicology animals at terminal necropsy.
Statistics:
Clinical observations were tabulated, but not statistically analyzed.

Body weight, body weight gain, food consumption and clinical pathology (haematology, coagulation and clinical chemistry) were analyzed for normality and equal variance. If the data set was normally distributed and of equal variance, statistical comparisons were conducted using a one-way analysis of variance (ANOVA), with post-hoc comparisons made using Dunnett's test. If normality and/or equal variance failed for a data set, statistical comparisons were conducted using the nonparametric Kruskal-Wallis ANOVA, with post-hoc comparisons made using Dunn's test. These parameters were compared using the statistical software provided by ToxData System, version 2.1.E.5. Urinalysis and organ weight data were compared using SYSTAT Software, version 10.2 (Systat Software Inc., Chicago, IL). Each sex was analyzed separately. Probability values of p < 0.05 (ToxData) or p 0.05 (SYSTAT) were considered significant.


Urinalysis (refractive index, specific gravity and pH only) and organ weight data were compared using ANOVA, with post-hoc comparisons made using Dunnett's test.

The Filtered Air Control group (Group 1) served as the control group for all comparisons.
Clinical signs:
no effects observed
Mortality:
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 examined
Water consumption and compound intake (if drinking water study):
not examined
Ophthalmological findings:
not examined
Haematological findings:
effects observed, treatment-related
Clinical biochemistry findings:
effects observed, treatment-related
Urinalysis findings:
effects observed, treatment-related
Behaviour (functional findings):
not examined
Organ weight findings including organ / body weight ratios:
effects observed, treatment-related
Gross pathological findings:
effects observed, treatment-related
Histopathological findings: non-neoplastic:
effects observed, treatment-related
Histopathological findings: neoplastic:
not examined
Details on results:
CLINICAL SIGNS AND MORTALITY
- No exposure-related deaths occurred during the study.
- The following clinical observations were observed during the pre- and post-exposure periods: skin/fur discoloration (blue), discoloration around the mouth, redness around nose fur, redness around the eyes, salivation, scab, injury, wet inguinal fur and limping. These observations (with the exception of skin/fur discoloration, discoloration around the mouth, scab, injury, and limping) are typical in nose-only exposure studies and were regarded as consequential to the rigors of nose-only exposure tube confinement.


BODY WEIGHT AND WEIGHT GAIN
- There were no statistically significant differences in group mean body weights for the male or female animals. Mean body weight gain was statistically significantly increased in High group females for Day 22-28, but the increase was not considered biologically relevant. No dose-related pattern was observed in either sex during the study.


FOOD CONSUMPTION
- There were no statistically significant differences in food consumption measurements, and no dose-related trend was observed in either sex.


HAEMATOLOGY
- At the end of the exposure period, white blood cells (WBC) and absolute eosinophils were statistically significantly increased in males in the Mid group, and absolute neutrophils, monocytes and eosinophils were statistically significantly increased in males in the High group, when compared to the Filtered Air Control. In females, relative reticulocytes were statistically significantly increased in the Mid group compared to the Filtered Air Control group. At the end of the recovery period, hemoglobin, hematocrit, MCV and absolute large unstained cells were statistically significantly increased in the High group males, when compared to the Filtered Air Control. At the end of the recovery period, absolute and relative reticulocytes were statistically significantly increased in the High group females, when compared to the Filtered Air Control. All red blood cell morphology observations were normal at the end of the exposure and recovery periods.
- Coagulation: At the end of the exposure period, no statistically significant differences were seen in males or females. No statistically significant differences were seen in males or females at the end of the recovery period.



CLINICAL CHEMISTRY
- At the end of the exposure period, calcium and glucose were statistically significantly increased in males in the Mid group and phosphorus was statistically significantly increased in Mid and High group males compared to the Filtered Air Control group. In females, gamma-glutamyl transpeptidase was significantly decreased in the Low group, calcium was statistically significantly increased in the Mid group, and chloride, phosphorus and albumin/globulin ratio levels were statistically significantly increased and globulin levels were statistically significantly decreased in the High group, compared to the Filtered Air Control group. At the end of the recovery period, cholesterol was statistically significantly increased in High group males, when compared to the Filtered Air Control. No statistically significant differences were seen in females at the end of the recovery period.


URINALYSIS
- Prior to terminal necropsy, urine refractive index and specific gravity were statistically significantly decreased for males in the low and High groups. At the end of the recovery period, no statistically significantly differences were observed between the groups.


ORGAN WEIGHTS
- At terminal necropsy, mean absolute and relative lung weights were statistically significantly increased in the Low, Mid and High groups for males, and in the Mid and High groups for females, compared to the Filtered Air Control group. At the recovery necropsy, mean absolute and relative lung weights were statistically significantly increased in males and females in the High group, mean relative testes weights were statistically significantly increased in males in the High group, and mean absolute and relative adrenal weights were statistically significantly decreased in males in the High group.


GROSS PATHOLOGY
- At Day 29, animals exposed to 0.65 mg/L air TBO had an increased incidence of pigmentation of the lungs which was correlated with the microscopic finding of alveolar foreign material and alveolar pigmented macrophages. Some animals exposed to 0.08 or 0.325 mg/L air levels also had pigmentation of the lungs with similar microscopic findings. In addition, the skin over the nasal bone was pigmented blue in all groups exposed to TBO. At Day 43, animals exposed to 0.65 mg/L air TBO had an increased incidence of pigmentation of the lungs which was correlated with the microscopic finding of alveolar foreign material and/or alveolar pigmented macrophages.


HISTOPATHOLOGY: NON-NEOPLASTIC
- At terminal sacrifice, there was an increase in the severity of alveolar pigmented macrophages, alveolar foreign material and individual alveolar foamy macrophages in animals exposed to all target concentrations of tungsten oxide. These three findings were considered related to exposure to TBO at all dose levels. In addition, at the 0.325 mg/L air and 0.65 mg/L air dose males there was an increase in the incidence of aggregates of foamy macrophages in the alveoli; this finding was also in the females exposed to 0.65 mg/L air concentration. There is a clear exposure response relationship in males for aggregates of macrophages in alveoli; this relationship is present, but less clear in the females. The finding of aggregated foamy macrophages was considered related to exposure of test substance at the 0.325 and 0.65 mg/L air concentrations in males and at the 0.65 mg/L air in females.

- After 14 days without exposure, there was the same incidence of animals with alveolar pigmented macrophages; however, the severity was slightly
decreased. There was a decrease in the severity of animals in Group 4 (0.65 mg/L air tungsten oxide) which had alveolar foreign material and individual alveolar foamy macrophages. There was a very small decrease in severity of foamy macrophage aggregates after 14 days without exposure. There was limited recovery of findings when compared to the terminal sacrifice animals.
Key result
Dose descriptor:
NOAEL
Effect level:
> 0.65 other: mg/L air (target)
Sex:
male/female
Basis for effect level:
other: No significant effects were observed at any dose
Key result
Dose descriptor:
NOAEL
Effect level:
> 0.652 mg/L air (analytical)
Sex:
male/female
Basis for effect level:
other: no significant effects were observed at any dose
Critical effects observed:
not specified

Overall means for tungsten oxide concentrations were determined gravimetrically to be 0.081, 0.331 and 0.652 mg/L for Groups 2 through 4, respectively. The tungsten oxide % recovery ranged from 100.83-102.38%. Small amounts of tungsten oxide in the chemically-analyzed filters for the Filtered Air Control group were attributed to contamination during the filter analysis processing and/or the calibration curve. The Filtered Air Control group filter-collected mean gravimetric value was 0.000 mg/L. The particle size distribution of the test atmosphere was within the respirable range. The overall mean tungsten oxide inhaled dose levels were 14.8, 60.2 and 118.8 mg/kg/day for Groups 2 through 4, respectively. The overall mean male tungsten oxide inhaled dose levels were 13.7, 55.7 and 110.2 mg/kg/day for Groups 2 through 4, respectively. The overall mean female tungsten oxide inhaled dose levels were 15.8, 64.7 and 127.3 mg/kg/day for Groups 2 through 4, respectively. The male inhaled dose levels were 10-11% below the target levels for all groups, while the female inhaled dose levels were 3-5% above the target levels for all groups. Prior to exposure initiation, the homogeneity of the test atmosphere in each tungsten oxide exposure chamber was confirmed.

Conclusions:
The NOAEL was determined to be > 0.65 mg/L exposed by nose-only tungsten oxide inhalation for 6 hours per day, 7 days per week for 28 consecutive days.
Executive summary:

No inhalation repeated dose toxicity data of sufficient quality are available for fused tungsten carbide (target substance). However, inhalation repeated dose toxicity data are available for tungsten oxide (source substance), which will be used for read-across. Due to similar water solubility and lower toxicity for the target substance compared to the source substance, the resulting read-across from the source substance to the target substance is appropriate. In addition, read-across is appropriate because the classification and labelling is similar for the source substance than the target substance, the PBT/vPvB profile is the same, and the dose descriptors are, or are expected to be, lower for the source substance. For more details, refer to the read-across category approach included in the Category section of this IUCLID submission and/or as an Annex in the CSR.

Endpoint conclusion
Endpoint conclusion:
no adverse effect observed
Dose descriptor:
NOAEC
650 mg/m³
Study duration:
subchronic
Species:
rat
Quality of whole database:
The reliability of this study for the substance tested is a K1, but in application of read-across to a different substance ECHA’s guidance specifies that the score can be a maximum of K2. Due to higher water solubility and greater in vitro bioaccessibility in synthetic alveolar, lysosomal, and interstitial fluids simulating inhalation exposure for the source substance, tungsten blue oxide (TBO) as compared to the target substance (tungsten metal) and lack of toxicity from acute toxicity studies for the target and source substances, toxicity data on the target substance is expected to represent a worse case, so read-across is appropriate between these substances. In addition, read-across is appropriate for this endpoint because the classification and labelling for human health toxicity endpoints is the same for the source and target substances, the PBT/vPvB profile is the same, and the dose descriptors are, or are expected to be, sufficiently similar or more conservative for the target substance. For more details, refer to the attached description of the read-across approach.

Repeated dose toxicity: inhalation - local effects

Endpoint conclusion
Endpoint conclusion:
no study available

Repeated dose toxicity: dermal - systemic effects

Endpoint conclusion
Endpoint conclusion:
no study available

Repeated dose toxicity: dermal - local effects

Endpoint conclusion
Endpoint conclusion:
no study available

Additional information

Repeated Dose Toxicity - Oral Route:

In addition to McCain et al (2015) rat oral 90-day repeated dose study, the US National Toxicology Program (NTP) has conducted two additional 90-day drinking water studies, one in Sprague-Dawley rats and a second one in B6C3F1 mice (10/sex/species/dose). The study design included doses of 0, 125, 250, 500, 1000, or 2000 mg/L. The in-life study phase has been completed but no study report has yet been issued. Currently, available in the US NTP website are graphs and Tables are preliminary results, but no full report has been issued. Furthermore, at the 2012 Annual Meeting of the Society of Toxicology, a Scientific Poster was presented detailing preliminary results of the NTP study. Preliminary results confirm the results of the McCain gavage study, showing the kidney as the major target organ for tungstate (especially at high drinking water doses of 1,000 and 2,000 mg/L). The US NTP’s Sodium Tungstate Dihydrate final study reports may be available in 2022.

On a rabbit 7-day maximum tolerable dose (MTD) study death was observed at 300 mg/kg/day and progressive weight loss at 200 mg/kg/day were reported suggesting that the oral MTD oral dose of sodium tungstate in the non-pregnant rabbit was less than 200 mg/kg/day.  Doses of 100 or 50 mg/kg/day were generally well tolerated with only small fluctuations in body weight. A high dose level between 100 and 200 mg/kg/day is concluded to be appropriate for the subsequent rabbit studies.

Repeated Dose Toxicity - Inhalation Route:

Repeated inhalation of 15 mg tungsten carbide dust/m3by rats for 13 weeks caused chronic rhinitis and mild histopathological alterations in the lung consisting of focal reactions around the terminal airways. The changes were characterized by minimal to moderate alveolar wall thickening, type II cell hyperplasia, and accumulations of pigmented macrophages. Mice exposed similarly to WC tolerated the treatment without symptoms except rhinitis in females (Kutzman RS and Drew RT. 1986).

Justification for classification or non-classification

Repeated Dose Toxicity - Oral Route:

No repeat dose toxicity data of sufficient quality were available for fused tungsten carbide; however, data were available for sodium tungstate, which will be used for reading-across. The LOAEL and NOAEL from the 90-day oral toxicity study were deemed to be 125 mg/kg/day and 75 mg/kg/day, respectively, based on histopathological effects reported in the kidneys of the 125 and 200 mg/kg/day dose groups. The BMDL10 derived from this data was calculated to be 102 mg/kg/day. The cutoff range for a category 2 classification under CLP for a 90-day oral toxicity study is between 10 and 100 mg/kg/day. The LOAEL of 125 mg/kg/day identified from the repeat dose oral toxicity study was greater than 100 mg/kg/day. In addition, the benchmark dose (BMDL10) based on the data from the 90-day oral toxicity studies, and using the kidney as the target organ, was calculated to be 102 mg/kg/day. Because the LOAEL from the 90-day study as well as the calculated BMDL10 were greater than the 100 mg/kg/day category 2 cutoff level under CLP, then a classification is not warranted.

Based on bioaccessibility studies of fused tungsten carbide that reported a 0.32% bioelution of tungsten after 5-hrs. incubation in gastric fluid, the amount of fused tungsten carbide needed to reach the dose to cause adverse effects is substantially higher than 1,000 mg/kg/day (limit dose). Therefore, no hazard classification is needed for fused tungsten carbide.

 

Repeated Dose Toxicity - Inhalation Route:

No repeat dose toxicity data of sufficient quality were available for fused tungsten carbide; however, data were available for tungsten carbide (WC) and tungsten oxide (also known as tungsten blue oxide or TBO), which will be used for read-across. No effects were reported in the 90 -day inhalation toxicity study on WC on which a classification could be based and the only dose tested was well below the classification criteria. Therefore, the dose from the 90-day inhalation toxicity study was insufficient for determining classification. Therefore, the 28-day inhalation toxicity study on TBO was used for read-across. The cutoff range for a category 2 classification under CLP for a 28-day inhalation toxicity study is between 0.06 and 0.6 mg/L. The NOAEL from the 28-day inhalation toxicity study on TBO was > 0.65 mg/L. Because the NOAEL from the 28-day study was greater than the 0.6 mg/L category 2 cutoff level, classification is not warranted.