Tungsten

Endpoint summary

Link to relevant study record(s)

Reference

Reference 1

Endpoint:

basic toxicokinetics in vitro / ex vivo

Type of information:

experimental study

Adequacy of study:

key study

Study period:

2009-01-28 To: 2009-05-01

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

test procedure in accordance with generally accepted scientific standards and described in sufficient detail

Remarks:

The bioaccessability study described in this report was performed according to the "Draft Guidance for RIP 3.6: Bioavailability and Read-Across for Metals and Minerals". GLP study.

Reason / purpose for cross-reference:

reference to other study

Objective of study:

absorption

Qualifier:

according to guideline

Guideline:

other: Draft Guidance for RIP 3.6: Bioavailability and Read-Across for Metals and Minerals

Deviations:

no

GLP compliance:

yes

Radiolabelling:

no

Species:

other: Human simulated fluids

Strain:

not specified

Sex:

not specified

Route of administration:

other: In vitro study

Details on exposure:

Preparation of Testing Fluids: The simulated fluids used in this study were gastric, lung alveolar, lung interstitial, lysosomal and artificial sweat. All salts and reagents used for the simulated fluid preparation were obtained from Sigma-Aldrich (St. Louis, MO).

Extraction Experiments: Extractions of the test substances in the simulated fluids were performed at pre-set time periods (up to 72 hours) while protected from light, using 0.1 g of sample in 50 ml of simulated fluid, at 37 °C and with continuous shaking (for all fluids except sweat, for which only initial shaking was performed). The experiments were carried out as follows:
- Simulated Gastric Fluid: The reactions were sampled for the determination of tungsten at 5 hours.
- Simulated Interstitial, Alveolar and Lysosomal Fluids: 5% CO2 in nitrogen was bubbled into solution at a rate of 50 ml/min. The reactions were sampled for the determination of tungsten at 2, 5, 24, and 72 hours.
- Simulated Sweat: The reactions were sampled for the determination of tungsten after 12 hours. No shaking was performed after the initial set up.

Sample Preparation: The simulated fluid extract samples were filtered immediately after sampling using 50 ml centrifuge tubes equipped with 0.45 microns PVDF filters (Grace Alltech, Deerfield, IL). The filtrates were stored in plastic bottles at 35 °C until analysis.

The extracts were then analyzed for tungsten by inductively coupled plasma-mass spectrometry (ICP-MS).

Duration and frequency of treatment / exposure:

Single application of tungsten with fluids. Simulated Gastric Fluid was sampled for the determination of tungsten at 5 hours. Simulated Interstitial, Alveolar and Lysosomal Fluids were sampled for the determination of tungsten at 2, 5, 24 and 72 hours. Simulated Sweat was sampled for the determination of tungsten after 12 hours.

Remarks:

Doses / Concentrations:
0.1 g of test substance in 50 mL of simulated fluid

Control animals:

no

Details on dosing and sampling:

PHARMACOKINETIC STUDY (Absorption, distribution, excretion)
- Tissues and body fluids sampled: Simulated gastric fluid, Simulated interstitial fluid, Simulated alveolar fluid, Simulated lysosomal fluid and Simulated sweat
- Time and frequency of sampling: Simulated gastric fluid sampled at 5 hours, Simulated interstitial fluid sampled at 2, 5, 24, and 72 hours, Simulated alveolar fluid sampled at 2, 5, 24 and 72 hours, Simulated lysosomal fluid sampled at 2, 5, 24 and 72 hours and Simulated sweat sampled after 12 hours.

Sample analysis: Samples were diluted (if necessary), spiked with an internal standard [bismuth (Bi) at 1,000 pg/mL, prepared from dilution of a 1,000 ug/mL Certified Standard; Ultra Scientific, North Kingston, RI and analyzed directly on a Perkin Elmer Elan DRC II ICP-MS equipped with a dynamic reaction cell (DRC) and PerkinElmer AS-93 Plus autosamples instrument, according to methods established at IITRI for this study. A standard curve (prepared from dilutions of a 10,000 5%HNO3/6% HF; inorganic Ventures, Lakewood, NJ] was analyzed along with samples on each day of analysis. Instruments calibrators were prepared by diluting Certified Standard with 0.5% nitric acid to concentrations of approximately 200; 400; 800; 1,600; 3,200; 6,400; 13,000 and 25,000 pg/mL.

Statistics:

Calibration curves, regression coefficients and r-squared values were calculated using PerkinElmer ICP-MS software and Microsoft Excel software. Concentration values of tungsten in the study samples were calculated from linear regression coefficients derived from calibration standards that bracketed the expected concentration levels of tungsten in the study samples.

Details on absorption:

The fluid extracts were diluted 1:2500 for analysis. The average amount of tungsten found in the extracts (expressed as percentage of the test material) was in the 0.054 to 0.97% range. The maximum solubility was determined at 72 hours for the simulated alveolar and lysosomal fluids (0.97 %). Percent relative standard deviations (%RSD; across all extraction times) ranged from 5.8 to 56%.

Gastric Fluid: The percent of available tungsten in simulated gastric fluid sampled at 5 hours was 0.23 +/- 0.013 % (5.8 % relative standard deviation).
Sweat Fluid: The percent of available tungsten in simulated sweat fluid sampled at 12 hours was 0.82 +/-0.14 % (18 % relative standard deviation).
Alveolar Fluid: The percent of available tungsten in simulated alveolar fluid sampled at 2, 5, 24, and 72 hours was 0.054 +/- 0.0044%, 0.072 +/- 0.0061%, 0.16 +/- 0.013%, and 0.97 +/- 0.13 %, respectively, with percent relative standard deviations of 8.0, 8.4, 8.0, and 13% RSD, respectively.
Lysosomal Fluid: The percent of available tungsten in simulated lysosomal fluid sampled at 2, 5, 24, and 72 hours was 0.20 +/- 0.072%, 0.32 +/- 0.16%, 0.48 +/- 0.18%, and 0.97+/- 0.38 %, respectively, with percent relative standard deviations of 36, 52, 38, and 39% RSD, respectively.
Interstitial Fluid: The percent of available tungsten in simulated interstitial fluid sampled at 2, 5, 24, and 72 hours was 0.079 +/- 0.016%, 0.090 +/- 0.032%, 0.17 +/- 0.096%, and 0.71 +/- 0.37 %, respectively, with percent relative standard deviations of 20, 36, 56, and 53% RSD, respectively.

Toxicokinetic parameters:

other: not applicable as this was an in vitro study

Metabolites identified:

not measured

Conclusions:

Interpretation of results (migrated information): bioaccumulation potential cannot be judged based on study results
The average amount of tungsten found in the extracts (expressed as percentage of the test material) was in the 0.054 to 0.97% range. The maximum solubility was determined at 72 hours for the simulated alveolar and lysosomal fluids (0.97 %). Percent relative standard deviations (%RSD; across all extraction times) ranged from 5.8 to 56%. In the simulated gastric fluid, the percent of available tungsten sampled at 5 hours was 0.23 +/- 0.013 % (5.8 % relative standard deviation). In the simulated sweat fluid, the percent of available tungsten sampled at 12 hours was 0.82 +/-0.14 % (18 % relative standard deviation).
In the simulated alveolar fluid, the percent of available tungsten sampled at 2, 5, 24 and 72 hours was 0.054 +/- 0.0044%, 0.072 +/- 0.0061%, 0.16 +/- 0.013%, and 0.97 +/- 0.13 %, respectively, with percent relative standard deviations of 8.0, 8.4, 8.0, and 13% RSD, respectively. In the simulated lysosomal fluid, the percent of available tungsten sampled at 2, 5, 24, and 72 hours was 0.20 +/- 0.072%, 0.32 +/- 0.16%, 0.48 +/- 0.18%, and 0.97 +/- 0.38 %, respectively, with percent relative standard deviations of 36, 52, 38, and 39% RSD, respectively. In the simulated interstitial fluid, the percent of available tungsten sampled at 2, 5, 24, and 72 hours was 0.079 +/- 0.016%, 0.090 +/- 0.032%, 0.17 +/- 0.096%, and 0.71 +/- 0.37 %, respectively, with percent relative standard deviations of 20, 36, 56, and 53% RSD, respectively. Based on the results, the bioavailability of tungsten metal would be expected to be low for the oral, dermal, and inhalation routes of administration.

Reference 2

Endpoint:

basic toxicokinetics

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, 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 on Annex 3 of the CSR.

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: Tungsten metal
3. CATEGORY APPROACH JUSTIFICATION: See Annex 3 in CSR
4. DATA MATRIX: See Annex 3 in CSR

Reason / purpose for cross-reference:

read-across: supporting information

Reason / purpose for cross-reference:

reference to same study

Objective of study:

toxicokinetics

Qualifier:

according to guideline

Guideline:

OECD Guideline 417 (Toxicokinetics)

Principles of method if other than guideline:

The experimental design included toxicokinetic (TK) animals for analysis of TBO content during the 28 days of exposure (TK Sets 1-4) and to assess the elimination phase of the test substance after one day of exposure (TK Set 5).

GLP compliance:

yes

Radiolabelling:

no

Species:

rat

Strain:

Sprague-Dawley

Sex:

male

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). 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) equipped with an automatic watering system.
-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 (eg 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 °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

Vehicle:

unchanged (no vehicle)

Details on 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 Aeorsol 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 analysed. 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

Duration and frequency of treatment / exposure:

Set 1: 6 hours/day, 7 days/week for 28 days
Set 2: 6 hours/day for 3 days
Set 3: 6 hours/day for 7 days
Set 4: 6 hours/day for 14 days
Set 5: 6 hours with 7 day recovery

Remarks:

Doses / Concentrations:
0.08 and 0.65 mg/L (Target tungsten oxide (aka tungsten blue oxide or TBO) concentration); 15.2 and 123.6 mg/kg/day (Target Inhaled Dose)

No. of animals per sex per dose / concentration:

Animals designated for toxicokinetic analysis were divided into five sets consisting of 4 males/set (time point) in each of the Low and High dose groups.

Control animals:

no

Positive control reference chemical:

no

Details on dosing and sampling:

PHARMACOKINETIC STUDY
- Tissues and body fluids sampled: at necropsy the brain, lungs, bone (femur), liver, kidney, spleen and reproductive organs were collected for analysis of TBO concentration. All toxicokinetic-designated animals were euthanized using sodium pentobarbital and exsanguinated from the abdominal aorta. Toxicokinetic animals were bled and necropsied as follows:

Set 1: Blood collection for the Low and High dose groups was performed on TK Day 0 [study Day 1 (pre-exposure)], and after the daily exposure on TK Days 1, 2, 3, 7, 14 and 28 (study Days 2, 3, 4, 8, 15 and 29); feces and urine were also collected from these animals overnight starting on TK Days 0, 1, 2, 3, 7, 14 and 28 (collection occurred the next day). These animals were necropsied on study Day 29. Blood was also collected on study Day 29, but was not chemically analysed.

Set 2: Blood collection for the Low and High dose groups was performed on TK Day 3 (study Day 4) after the daily exposure and preserved for future evaluation. These animals were necropsied on TK Day 3.

Set 3: Blood collection for the Low and High dose groups was performed on TK Day 7 (study Day 8) after the daily exposure and preserved for future evaluation. These animals were necropsied on TK Day 7.

Set 4: Blood collection for the Low and High dose groups was performed on TK Day 14 (study Day 15) after the daily exposure and preserved for future evaluation. These animals were necropsied on TK Day 14.

Set 5: Blood collection for the Low and High dose groups was performed on this group on TK Day 0 [study Day 28 (pre-exposure)], after the exposure on TK Day 1 (study Day 29), and recovery Days 2, 3, 4, 5, 6 and 7) (study Days 30, 31, 32, 33, 34 and 35); feces and urine were also collected from these animals overnight starting on TK Days 0, 1, 2, 3, 4, 5, 6 and 7 (collection occurred the next day). These animals were necropsied on TK Day 8 (study Day 36) after the final urine and feces collection. Blood was also collected on TK Day 8, but was not chemically analysed.

Statistics:

no data

Details on absorption:

In the single dose experiment, tungsten was absorbed systemically and blood tungsten concentration reached maximum (Tmax) at 0 hours post-exposure.

Details on distribution in tissues:

Repeat dose experiment:

From the repeat dose experiments, tungsten concentration in lung tissue was at least one order of magnitude greater than in any other organ collected, the lung being the primary organ for TBO exposure via inhalation. In general, the femur and kidney ranked second and third highest in tungsten concentration on a per organ weight basis, respectively. The results indicated that the lung, femur and liver ranked first, second and third for total tungsten burden, respectively. The study day appeared to have little or no effect on the rank of tungsten distribution in tissue and organs. Tungsten concentration in tissue and organs increased with increasing dose, but not in a dose-proportionate manner.
Tungsten concentration in each organ increased with increasing inhalation exposure time for the lung, liver, kidney, spleen, testes, brain and femur, respectively. The increment was not significantly different between study Days 7 and 14, suggesting that steady-state absorption of inhaled TBO was reached on study Day 14. Compared with the steady-state concentration on study Day 14, tungsten in liver, kidney, testes and brain decreased at least 50% on study Day 29; however, unlike toxicokinetic Set 2 to 4 animals (necropsied immediately following termination of inhalation exposure on study Day 3, 7 or 14, respectively), the Set 1 animals were necropsied 24 hours after termination of exposure on study Day 29 to allow for overnight urine and feces collection. The results suggest that the functional elimination half-life of tungsten in those organs was less than 24 hours. On the contrary, tungsten in femur on study Day 29 was no different as compared to study Day 14, indicating that elimination of tungsten from femur was relatively slow as compared to other organs. The results were consistent with the Set 5 results (animals necropsied on Recovery Day 8 following a single inhalation exposure). Tungsten in femur on Recovery Day 8 (Set 5) was approximately one third as compared to that on study Day 3 (Set 2 animals) following multiple inhalation exposure, while tungsten concentration in the rest of organs was at least one order of magnitude lower.

Single administration experiment:

Following inhalation exposure of tungsten oxide (aka tungsten blue oxide or TBO), tungsten was absorbed systemically, as indicated by maximum blood concentration at 0 hour post-exposure. At 48 hours post-exposure, there was a slow elimination phase following an initial fast elimination phase for the 0.65 mg/L dose group (High). However, the second slow elimination phase was not evident for the 0.08 mg/L dose group (Low), which may be due to background noise of tungsten concentration. Based on the concentrations in the blood, the terminal phase half-life values were determined to be 23 ± 4.3 and 154 ± 92.8 hours for the 0.08 and 0.65 mg/L dose groups, respectively. Maximum blood tungsten concentration was 0.819 ± 0.215 and 10.9 ± 4.7 1.1g/g and area under the blood concentration-time curve extrapolated to infinity was 11.8 ± 3.29 and 148 ± 33.7 hr*i.tg/g for the 0.08 and 0.65 mg/L dose groups, respectively. These two exposure parameters increased proportionally with increasing TBO dose level. The systemic clearance based on the concentrations in the blood was 1.24 ± 0.39 and 0.78 ± 0.18 L/hr/kg for the 0.08 and 0.65 mg/L dose groups, respectively.

Details on excretion:

Repeat dose experiment:

The results from the repeat dose experiments indicate that the amount of tungsten excreted from feces was approximately three orders of magnitude greater than from urine. Since the nasal cavities are the major deposition site for inhaled particles in the rate, the study results suggest that most of the inhaled tungsten oxide (aka tungsten blue oxide or TBO) was deposited in the nasal passages and subsequently ingested into the gastrointestinal tract and excreted with the feces. The study results also indicate that the excretion rate of tungsten from feces and urine and tungsten concentration in blood increased with increasing TBO dose level; however, the effect of study day was sporadic and a discernible trend was not observed.

Single administration experiment:

From the single dose experiment, the results indicate that excretion of tungsten from the gastrointestinal tract was negligible after post-exposure Day 2 and 3 for Groups 2 (Low) and 4 (High), respectively. The results are consistent with the general consensus that insoluble particles deposited in the nasal and tracheobronchial airways would be cleared within 24 hours through mucus escalator followed by ingestion into gastrointestinal tract and excretion with the feces. In contrast, there was still detectable tungsten in urine on post-exposure Day 7.

Toxicokinetic parameters:

AUC: 11.8 ± 3.29 hr ug/g for the 0.08 mg/L dose group

Toxicokinetic parameters:

Cmax: 0.819 ± 0.215 ug/g for the 0.08 mg/L dose group

Toxicokinetic parameters:

half-life 1st: 23 ± 4.3 hours for the 0.08 mg/L dose group

Toxicokinetic parameters:

AUC: 148 ± 33.7 hr ug/g for the 0.65 mg/L dose group

Toxicokinetic parameters:

Cmax: 10.9 ± 4.7 ug/g for the 0.65 mg/L dose group

Toxicokinetic parameters:

half-life 1st: 154 ± 92.8 hours for the 0.65 mg/L dose group

Metabolites identified:

not measured

The mean MMADs of the test atmosphere were 2.63 and 2.74 um with GSDs of 1.89 and 1.92 for the low and high dose groups, respectively.

Overall means for tungsten oxide (aka tungsten blue oxide or TBO) concentrations were determined gravimetrically to be 0.081and 0.652 mg/L for the low and high dose groups, respectively. The TBO % recovery ranged from 100.83-102.38%. Small amounts of TBO 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 TBO inhaled dose levels were 14.8 and 118.8 mg/kg/day for the lowand high dose groups, respectively. The overall mean male TBO inhaled dose levels were 13.7 and 110.2 mg/kg/day for the low and high dose groups, respectively. The overall mean female TBO inhaled dose levels were 15.8 and 127.3 mg/kg/day for the low and high dose groups, 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 TBO exposure chamber was confirmed.

Conclusions:

During the toxicokinetic analysis, two phases were explored through the data analysis of the tungsten oxide (aka tungsten blue oxide or TBO) present in the bio-samples: the accumulation
phase and the elimination phase. The accumulation phase encompassed the data from Sets 1, 2, 3 and 4. The objectives were: 1) to determine what major organs/tissues had TBO distributed in them; 2) to determine the uptake profiles of TBO in major organs/tissues over time; 3) to determine whether TBO reached a steady- state or accumulated in any organs/tissues for a 28-day dosing period; and 4) to determine the total body burden of TBO over time. The data treatment for Sets 1, 2, 3 and 4 included the tissue and organ distribution of TBO plotted as the percentage of inhaled dose, the percentage of inhaled dose per unit weight of the organs/tissues and the composite mean of TBO body burden (ie total TBO recovered from each animal as the percentage of daily inhaled dose per unit body weight) versus time profile. The elimination phase encompassed the data from Set 5. The objectives were 1) to determine the TBO elimination rate, its half-life and clearance rate; 2) to determine what major organs/tissues had TBO distributed in them after a 7-day clearance; and 3) to determine the total body burden of TBO after a 7-day clearance.

The data treatment for Set 5 included 1) non-compartmental pharmacokinetic analysis on the whole blood, urine and feces TBO concentration versus time profiles from the Set 5 animals using WinNonlin software, version 4.1 (Pharsight Corp, Mountain View, CA); 2) tissue and organ distribution of TBO from Set 5 plotted as the percentage of inhaled dose; and 3) composite mean of TBO body burden (i.e., total TBO recovered from each animal as the percentage of daily inhaled dose per unit body weight) from Set 5.
Following inhalation exposure to TBO, tungsten was absorbed systemically with blood tungsten concentrations reaching maximum at 0 hours post-exposure. The exposure parameters of Cmax and AUC increased proportionally with increasing TBO dose level: Cmax was 0.819 ± 0.215 and 10.9 ± 4.7 fig/g and AUC was 11.8 ± 3.29 and 148 ± 33.7 hr*Rg/g for the 0.08 and 0.65 mg/L dose groups, respectively. The systemic elimination half-life was 23 ± 4.3 and 154 ± 92.8 hours for the 0.08 and 0.65 mg/L dose groups, respectively, and the clearance rate was 1.24 ± 0.39 and 0.78 ± 0.18 L/hr/kg for the 0.08 and 0.65 mg/L dose groups, respectively. The majority of the inhaled TBO was excreted through the gastrointestinal tract.
Tungsten concentration in lung tissue was at least one order of magnitude greater than in any other organ collected. In general, the femur and kidney ranked second and third highest, respectively, in tungsten concentration on a per organ weight basis. In terms of total tungsten burden in the organs, the lung, femur and liver ranked first, second and third, respectively. Tungsten distribution in tissue and organs reached steady-state on study Day 14. With the exception of the femur, the functional elimination half-life of tungsten in most organs was less than 24 hours.

Executive summary:

No inhalation toxicokinetic data of sufficient quality are available for tungsten (target substance). However, data are available for tungsten oxide (source substance), which are 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 on Annex 3 in the CSR. 

Reference 3

Endpoint:

basic toxicokinetics in vitro / ex vivo

Type of information:

experimental study

Adequacy of study:

key study

Study period:

2009-01-28 To: 2009-05-01

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

test procedure in accordance with generally accepted scientific standards and described in sufficient detail

Remarks:

The bioaccessability study described in this report was performed according to the "Draft Guidance for RIP 3.6: Bioavailability and Read-Across for Metals and Minerals". GLP study.

Reason / purpose for cross-reference:

reference to other study

Objective of study:

absorption

Qualifier:

according to guideline

Guideline:

other: Draft Guidance for RIP 3.6: Bioavailability and Read-Across for Metals and Minerals

Deviations:

no

GLP compliance:

yes

Radiolabelling:

no

Species:

other: Human simulated fluids

Strain:

not specified

Sex:

not specified

Route of administration:

other: In vitro study

Details on exposure:

Preparation of Testing Fluids: The simulated fluids used in this study were gastric, lung alveolar, lung interstitial, lysosomal and artificial sweat. All salts and reagents used for the simulated fluid preparation were obtained from Sigma-Aldrich (St. Louis, MO).

Extraction Experiments: Extractions of the test substances in the simulated fluids were performed at pre-set time periods (up to 72 hours) while protected from light, using 0.1 g of sample in 50 ml of simulated fluid, at 37 °C and with continuous shaking (for all fluids except sweat, for which only initial shaking was performed). The experiments were carried out as follows:
- Simulated Gastric Fluid: The reactions were sampled for the determination of tungsten at 5 hours.
- Simulated Interstitial, Alveolar and Lysosomal Fluids: 5% CO2 in nitrogen was bubbled into solution at a rate of 50 ml/min. The reactions were sampled for the determination of tungsten at 2, 5, 24 and 72 hours.
- Simulated Sweat: The reactions were sampled for the determination of tungsten after 12 hours. No shaking was performed after the initial set up.

Sample Preparation: The simulated fluid extract samples were filtered immediately after sampling using 50 ml centrifuge tubes equipped with 0.45 microns PVDF filters (Grace Alltech, Deerfield, IL). The filtrates were stored in plastic bottles at 35 °C until analysis.

The extracts were then analyzed for tungsten by inductively coupled plasma-mass spectrometry (ICP-MS).

Duration and frequency of treatment / exposure:

Single application of tungsten with fluids. Simulated Gastric Fluid was sampled for the determination of tungsten at 5 hours. Simulated Interstitial, Alveolar and Lysosomal Fluids were sampled for the determination of tungsten at 2, 5, 24 and 72 hours. Simulated Sweat was sampled for the determination of tungsten after 12 hours.

Remarks:

Doses / Concentrations:
0.1 g of test substance in 50 mL of simulated fluid

Control animals:

no

Details on dosing and sampling:

PHARMACOKINETIC STUDY (Absorption, distribution, excretion)
- Tissues and body fluids sampled: Simulated gastric fluid, Simulated interstitial fluid, Simulated alveolar fluid, Simulated lysosomal fluid and Simulated sweat
- Time and frequency of sampling: Simulated gastric fluid sampled at 5 hours, Simulated interstitial fluid sampled at 2, 5, 24 and 72 hours, Simulated alveolar fluid sampled at 2, 5, 24 and 72 hours, Simulated lysosomal fluid sampled at 2, 5, 24 and 72 hours and Simulated sweat sampled after 12 hours.

Sample analysis: Samples were diluted (if necessary), spiked with an internal standard [bismuth (Bi) at 1,000 pg/mL, prepared from dilution of a 1,000 ug/mL Certified Standard; Ultra Scientific, North Kingston, RI and analysed directly on a Perkin Elmer Elan DRC II ICP-MS equipped with a dynamic reaction cell (DRC) and PerkinElmer AS-93 Plus autosamples instrument, according to methods established at IITRI for this study. A standard curve (prepared from dilutions of a 10,000 5%HNO3/6% HF; inorganic Ventures, Lakewood, NJ] was analyzed along with samples on each day of analysis. Instruments calibrators were prepared by diluting Certified Standard with 0.5% nitric acid to concentrations of approximately 200; 400; 800; 1,600; 3,200; 6,400; 13,000 and 25,000 pg/mL.

Statistics:

Calibration curves, regression coefficients and r-squared values were calculated using PerkinElmer ICP-MS software and Microsoft Excel software. Concentration values of tungsten in the study samples were calculated from linear regression coefficients derived from calibration standards that bracketed the expected concentration levels of tungsten in the study samples.

Details on absorption:

The fluid extracts were diluted 1:2500 for analysis. The average amount of tungsten found in the extracts (expressed as percentage of the test material) was in the 0.054 to 0.97% range. The maximum solubility was determined at 72 hours for the simulated alveolar (0.97 %). Percent relative standard deviations (%RSD; across all extraction times) ranged from 5.8 to 56%.
Alveolar Fluid: The percent of available tungsten in simulated alveolar fluid sampled at 2, 5, 24 and 72 hours was 0.054 +/- 0.0044%, 0.072 +/- 0.0061%, 0.16 +/- 0.013% and 0.97 +/- 0.13 %, respectively, with percent relative standard deviations of 8.0, 8.4, 8.0 and 13% RSD, respectively.

Toxicokinetic parameters:

other: not applicable as this was an in vitro study

Metabolites identified:

not measured

Conclusions:

Interpretation of results (migrated information): bioaccumulation potential cannot be judged based on study results
The average amount of tungsten found in the extracts (expressed as percentage of the test material) was in the 0.054 to 0.97% range. The maximum solubility was determined at 72 hours for the simulated alveolar (0.97 %). Percent relative standard deviations (%RSD; across all extraction times) ranged from 5.8 to 56%.
In the simulated alveolar fluid, the percent of available tungsten sampled at 2, 5, 24 and 72 hours was 0.054 +/- 0.0044%, 0.072 +/- 0.0061%, 0.16 +/- 0.013% and 0.97 +/- 0.13 %, respectively, with percent relative standard deviations of 8.0, 8.4, 8.0 and 13% RSD, respectively. Based on the results, the bioavailability of tungsten metal would be expected to be low for the oral, dermal and inhalation routes of administration.

Executive summary:

Bioaccumulation potential cannot be judged based on study results.

Reference 4

Endpoint:

basic toxicokinetics in vitro / ex vivo

Type of information:

experimental study

Adequacy of study:

key study

Study period:

2009-01-28 To: 2009-05-01

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

test procedure in accordance with generally accepted scientific standards and described in sufficient detail

Remarks:

The bioaccessability study described in this report was performed according to the "Draft Guidance for RIP 3.6: Bioavailability and Read-Across for Metals and Minerals". GLP study.

Reason / purpose for cross-reference:

reference to other study

Objective of study:

absorption

Qualifier:

according to guideline

Guideline:

other: Draft Guidance for RIP 3.6: Bioavailability and Read-Across for Metals and Minerals

Deviations:

no

GLP compliance:

yes

Radiolabelling:

no

Species:

other: Human simulated fluids

Strain:

not specified

Sex:

not specified

Route of administration:

other: In vitro study

Details on exposure:

Preparation of Testing Fluids: The simulated fluids used in this study were gastric, lung alveolar, lung interstitial, lysosomal and artificial sweat. All salts and reagents used for the simulated fluid preparation were obtained from Sigma-Aldrich (St. Louis, MO).

Extraction Experiments: Extractions of the test substances in the simulated fluids were performed at pre-set time periods (up to 72 hours) while protected from light, using 0.1 g of sample in 50 ml of simulated fluid, at 37 °C and with continuous shaking (for all fluids except sweat, for which only initial shaking was performed). The experiments were carried out as follows:
- Simulated Gastric Fluid: The reactions were sampled for the determination of tungsten at 5 hours.
- Simulated Interstitial, Alveolar and Lysosomal Fluids: 5% CO2 in nitrogen was bubbled into solution at a rate of 50 ml/min. The reactions were sampled for the determination of tungsten at 2, 5, 24 and 72 hours.
- Simulated Sweat: The reactions were sampled for the determination of tungsten after 12 hours. No shaking was performed after the initial set up.

Sample Preparation: The simulated fluid extract samples were filtered immediately after sampling using 50 ml centrifuge tubes equipped with 0.45 microns PVDF filters (Grace Alltech, Deerfield, IL). The filtrates were stored in plastic bottles at 35 °C until analysis.

The extracts were then analysed for tungsten by inductively coupled plasma-mass spectrometry (ICP-MS).

Duration and frequency of treatment / exposure:

Single application of tungsten with fluids. Simulated Gastric Fluid was sampled for the determination of tungsten at 5 hours. Simulated Interstitial, Alveolar and Lysosomal Fluids were sampled for the determination of tungsten at 2, 5, 24 and 72 hours. Simulated Sweat was sampled for the determination of tungsten after 12 hours.

Remarks:

Doses / Concentrations:
0.1 g of test substance in 50 mL of simulated fluid

Control animals:

no

Details on dosing and sampling:

PHARMACOKINETIC STUDY (Absorption, distribution, excretion)
- Tissues and body fluids sampled: Simulated gastric fluid, Simulated interstitial fluid, Simulated alveolar fluid, Simulated lysosomal fluid and Simulated sweat
- Time and frequency of sampling: Simulated gastric fluid sampled at 5 hours, Simulated interstitial fluid sampled at 2, 5, 24, and 72 hours, Simulated alveolar fluid sampled at 2, 5, 24 and 72 hours, Simulated lysosomal fluid sampled at 2, 5, 24 and 72 hours and Simulated sweat sampled after 12 hours.

Sample analysis: Samples were diluted (if necessary), spiked with an internal standard [bismuth (Bi) at 1,000 pg/mL, prepared from dilution of a 1,000 ug/mL Certified Standard; Ultra Scientific, North Kingston, RI and analysed directly on a Perkin Elmer Elan DRC II ICP-MS equipped with a dynamic reaction cell (DRC) and PerkinElmer AS-93 Plus autosamples instrument, according to methods established at IITRI for this study. A standard curve (prepared from dilutions of a 10,000 5%HNO3/6% HF; inorganic Ventures, Lakewood, NJ] was analyzed along with samples on each day of analysis. Instruments calibrators were prepared by diluting Certified Standard with 0.5% nitric acid to concentrations of approximately 200; 400; 800; 1,600; 3,200; 6,400; 13,000 and 25,000 pg/mL.

Statistics:

Calibration curves, regression coefficients and r-squared values were calculated using PerkinElmer ICP-MS software and Microsoft Excel software. Concentration values of tungsten in the study samples were calculated from linear regression coefficients derived from calibration standards that bracketed the expected concentration levels of tungsten in the study samples.

Details on absorption:

The fluid extracts were diluted 1:2500 for analysis. The average amount of tungsten found in the extracts (expressed as percentage of the test material) was in the 0.054 to 0.97% range. Percent relative standard deviations (%RSD; across all extraction times) ranged from 5.8 to 56%. Gastric Fluid: The percent of available tungsten in simulated gastric fluid sampled at 5 hours was 0.23 +/- 0.013 % (5.8 % relative standard deviation).

Toxicokinetic parameters:

other: not applicable as this was an in vitro study

Metabolites identified:

not measured

Conclusions:

Interpretation of results (migrated information): bioaccumulation potential cannot be judged based on study results
The average amount of tungsten found in the extracts (expressed as percentage of the test material) was in the 0.054 to 0.97% range. Percent relative standard deviations (%RSD; across all extraction times) ranged from 5.8 to 56%. In the simulated gastric fluid, the percent of available tungsten sampled at 5 hours was 0.23 +/- 0.013 % (5.8 % relative standard deviation). In the simulated sweat fluid, the percent of available tungsten sampled at 12 hours was 0.82 +/-0.14 % (18 % relative standard deviation). Based on the results, the bioavailability of tungsten metal would be expected to be low for the oral, dermal and inhalation routes of administration.

Executive summary:

Bioaccumulation potential cannot be judged based on study results.

Reference 5

Endpoint:

basic toxicokinetics in vitro / ex vivo

Type of information:

experimental study

Adequacy of study:

key study

Study period:

2009-01-28 To: 2009-05-01

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

test procedure in accordance with generally accepted scientific standards and described in sufficient detail

Remarks:

The bioaccessability study described in this report was performed according to the "Draft Guidance for RIP 3.6: Bioavailability and Read-Across for Metals and Minerals". GLP study.

Reason / purpose for cross-reference:

reference to other study

Objective of study:

absorption

Qualifier:

according to guideline

Guideline:

other: Draft Guidance for RIP 3.6: Bioavailability and Read-Across for Metals and Minerals

Deviations:

no

GLP compliance:

yes

Radiolabelling:

no

Species:

other: Human simulated fluids

Strain:

not specified

Sex:

not specified

Route of administration:

other: In vitro study

Details on exposure:

Preparation of Testing Fluids: The simulated fluids used in this study were gastric, lung alveolar, lung interstitial, lysosomal and artificial sweat. All salts and reagents used for the simulated fluid preparation were obtained from Sigma-Aldrich (St. Louis, MO).

Extraction Experiments: Extractions of the test substances in the simulated fluids were performed at pre-set time periods (up to 72 hours) while protected from light, using 0.1 g of sample in 50 ml of simulated fluid, at 37 °C and with continuous shaking (for all fluids except sweat, for which only initial shaking was performed). The experiments were carried out as follows:
- Simulated Gastric Fluid: The reactions were sampled for the determination of tungsten at 5 hours.
- Simulated Interstitial, Alveolar and Lysosomal Fluids: 5% CO2 in nitrogen was bubbled into solution at a rate of 50 ml/min. The reactions were sampled for the determination of tungsten at 2, 5, 24, and 72 hours.
- Simulated Sweat: The reactions were sampled for the determination of tungsten after 12 hours. No shaking was performed after the initial set up.

Sample Preparation: The simulated fluid extract samples were filtered immediately after sampling using 50 ml centrifuge tubes equipped with 0.45 microns PVDF filters (Grace Alltech, Deerfield, IL). The filtrates were stored in plastic bottles at 35 °C until analysis.

The extracts were then analysed for tungsten by inductively coupled plasma-mass spectrometry (ICP-MS).

Duration and frequency of treatment / exposure:

Single application of tungsten with fluids. Simulated Gastric Fluid was sampled for the determination of tungsten at 5 hours. Simulated Interstitial, Alveolar and Lysosomal Fluids were sampled for the determination of tungsten at 2, 5, 24 and 72 hours. Simulated Sweat was sampled for the determination of tungsten after 12 hours.

Remarks:

Doses / Concentrations:
0.1 g of test substance in 50 mL of simulated fluid

Control animals:

no

Details on dosing and sampling:

PHARMACOKINETIC STUDY (Absorption, distribution, excretion)
- Tissues and body fluids sampled: Simulated gastric fluid, Simulated interstitial fluid, Simulated alveolar fluid, Simulated lysosomal fluid and Simulated sweat
- Time and frequency of sampling: Simulated gastric fluid sampled at 5 hours, Simulated interstitial fluid sampled at 2, 5, 24 and 72 hours, Simulated alveolar fluid sampled at 2, 5, 24,and 72 hours, Simulated lysosomal fluid sampled at 2, 5, 24 and 72 hours and Simulated sweat sampled after 12 hours.

Sample analysis: Samples were diluted (if necessary), spiked with an internal standard [bismuth (Bi) at 1,000 pg/mL, prepared from dilution of a 1,000 ug/mL Certified Standard; Ultra Scientific, North Kingston, RI and analysed directly on a Perkin Elmer Elan DRC II ICP-MS equipped with a dynamic reaction cell (DRC) and PerkinElmer AS-93 Plus autosamples instrument, according to methods established at IITRI for this study. A standard curve (prepared from dilutions of a 10,000 5%HNO3/6% HF; inorganic Ventures, Lakewood, NJ] was analyzed along with samples on each day of analysis. Instruments calibrators were prepared by diluting Certified Standard with 0.5% nitric acid to concentrations of approximately 200; 400; 800; 1,600; 3,200; 6,400; 13,000 and 25,000 pg/mL.

Statistics:

Calibration curves, regression coefficients and r-squared values were calculated using PerkinElmer ICP-MS software and Microsoft Excel software. Concentration values of tungsten in the study samples were calculated from linear regression coefficients derived from calibration standards that bracketed the expected concentration levels of tungsten in the study samples.

Details on absorption:

The fluid extracts were diluted 1:2500 for analysis. The average amount of tungsten found in the extracts (expressed as percentage of the test material) was in the 0.054 to 0.97% range.Percent relative standard deviations (%RSD; across all extraction times) ranged from 5.8 to 56%.
Interstitial Fluid: The percent of available tungsten in simulated interstitial fluid sampled at 2, 5, 24 and 72 hours was 0.079 +/- 0.016%, 0.090 +/- 0.032%, 0.17 +/- 0.096% and 0.71 +/- 0.37 %, respectively, with percent relative standard deviations of 20, 36, 56 and 53% RSD, respectively.

Toxicokinetic parameters:

other: not applicable as this was an in vitro study

Metabolites identified:

not measured

Conclusions:

Interpretation of results (migrated information): bioaccumulation potential cannot be judged based on study results
The average amount of tungsten found in the extracts (expressed as percentage of the test material) was in the 0.054 to 0.97% range. Percent relative standard deviations (%RSD; across all extraction times) ranged from 5.8 to 56%. In the simulated interstitial fluid, the percent of available tungsten sampled at 2, 5, 24 and 72 hours was 0.079 +/- 0.016%, 0.090 +/- 0.032%, 0.17 +/- 0.096% and 0.71 +/- 0.37 %, respectively, with percent relative standard deviations of 20, 36, 56 and 53% RSD, respectively. Based on the results, the bioavailability of tungsten metal would be expected to be low for the oral, dermal, and inhalation routes of administration.

Executive summary:

Bioaccumulation potential cannot be judged based on study results.

Reference 6

Endpoint:

basic toxicokinetics in vitro / ex vivo

Type of information:

experimental study

Adequacy of study:

key study

Study period:

2009-01-28 To: 2009-05-01

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

test procedure in accordance with generally accepted scientific standards and described in sufficient detail

Remarks:

The bioaccessability study described in this report was performed according to the "Draft Guidance for RIP 3.6: Bioavailability and Read-Across for Metals and Minerals". GLP study.

Reason / purpose for cross-reference:

reference to other study

Objective of study:

absorption

Qualifier:

according to guideline

Guideline:

other: Draft Guidance for RIP 3.6: Bioavailability and Read-Across for Metals and Minerals

Deviations:

no

GLP compliance:

yes

Radiolabelling:

no

Species:

other: Human simulated fluids

Strain:

not specified

Sex:

not specified

Route of administration:

other: In vitro study

Details on exposure:

Preparation of Testing Fluids: The simulated fluids used in this study were gastric, lung alveolar, lung interstitial, lysosomal and artificial sweat. All salts and reagents used for the simulated fluid preparation were obtained from Sigma-Aldrich (St. Louis, MO).

Extraction Experiments: Extractions of the test substances in the simulated fluids were performed at pre-set time periods (up to 72 hours) while protected from light, using 0.1 g of sample in 50 ml of simulated fluid, at 37 °C and with continuous shaking (for all fluids except sweat, for which only initial shaking was performed). The experiments were carried out as follows:
- Simulated Gastric Fluid: The reactions were sampled for the determination of tungsten at 5 hours.
- Simulated Interstitial, Alveolar and Lysosomal Fluids: 5% CO2 in nitrogen was bubbled into solution at a rate of 50 ml/min. The reactions were sampled for the determination of tungsten at 2, 5, 24 and 72 hours.
- Simulated Sweat: The reactions were sampled for the determination of tungsten after 12 hours. No shaking was performed after the initial set up.

Sample Preparation: The simulated fluid extract samples were filtered immediately after sampling using 50 ml centrifuge tubes equipped with 0.45 microns PVDF filters (Grace Alltech, Deerfield, IL). The filtrates were stored in plastic bottles at 35 °C until analysis.

The extracts were then analysed for tungsten by inductively coupled plasma-mass spectrometry (ICP-MS).

Duration and frequency of treatment / exposure:

Single application of tungsten with fluids. Simulated Gastric Fluid was sampled for the determination of tungsten at 5 hours. Simulated Interstitial, Alveolar and Lysosomal Fluids were sampled for the determination of tungsten at 2, 5, 24, and 72 hours. Simulated Sweat was sampled for the determination of tungsten after 12 hours.

Remarks:

Doses / Concentrations:
0.1 g of test substance in 50 mL of simulated fluid

Control animals:

no

Details on dosing and sampling:

PHARMACOKINETIC STUDY (Absorption, distribution, excretion)
- Tissues and body fluids sampled: Simulated gastric fluid, Simulated interstitial fluid, Simulated alveolar fluid, Simulated lysosomal fluid and Simulated sweat
- Time and frequency of sampling: Simulated gastric fluid sampled at 5 hours, Simulated interstitial fluid sampled at 2, 5, 24, and 72 hours, Simulated alveolar fluid sampled at 2, 5, 24 and 72 hours, Simulated lysosomal fluid sampled at 2, 5, 24 and 72 hours and Simulated sweat sampled after 12 hours.

Sample analysis: Samples were diluted (if necessary), spiked with an internal standard [bismuth (Bi) at 1,000 pg/mL, prepared from dilution of a 1,000 ug/mL Certified Standard; Ultra Scientific, North Kingston, RI and analyzed directly on a Perkin Elmer Elan DRC II ICP-MS equipped with a dynamic reaction cell (DRC) and PerkinElmer AS-93 Plus autosamples instrument, according to methods established at IITRI for this study. A standard curve (prepared from dilutions of a 10,000 5%HNO3/6% HF; inorganic Ventures, Lakewood, NJ] was analysed along with samples on each day of analysis. Instruments calibrators were prepared by diluting Certified Standard with 0.5% nitric acid to concentrations of approximately 200; 400; 800; 1,600; 3,200; 6,400; 13,000 and 25,000 pg/mL.

Statistics:

Calibration curves, regression coefficients and r-squared values were calculated using PerkinElmer ICP-MS software and Microsoft Excel software. Concentration values of tungsten in the study samples were calculated from linear regression coefficients derived from calibration standards that bracketed the expected concentration levels of tungsten in the study samples.

Details on absorption:

The fluid extracts were diluted 1:2500 for analysis. The average amount of tungsten found in the extracts (expressed as percentage of the test material) was in the 0.054 to 0.97% range. The maximum solubility was determined at 72 hours for the lysosomal fluids (0.97 %). Percent relative standard deviations (%RSD; across all extraction times) ranged from 5.8 to 56%.
Lysosomal Fluid: The percent of available tungsten in simulated lysosomal fluid sampled at 2, 5, 24 and 72 hours was 0.20 +/- 0.072%, 0.32 +/- 0.16%, 0.48 +/- 0.18% and 0.97+/- 0.38 %, respectively, with percent relative standard deviations of 36, 52, 38 and 39% RSD, respectively.

Toxicokinetic parameters:

other: not applicable as this was an in vitro study

Metabolites identified:

not measured

Conclusions:

Interpretation of results (migrated information): bioaccumulation potential cannot be judged based on study results
The average amount of tungsten found in the extracts (expressed as percentage of the test material) was in the 0.054 to 0.97% range. The maximum solubility was determined at 72 hours for the lysosomal fluids (0.97 %). Percent relative standard deviations (%RSD; across all extraction times) ranged from 5.8 to 56%. In the simulated lysosomal fluid, the percent of available tungsten sampled at 2, 5, 24 and 72 hours was 0.20 +/- 0.072%, 0.32 +/- 0.16%, 0.48 +/- 0.18% and 0.97 +/- 0.38 %, respectively, with percent relative standard deviations of 36, 52, 38 and 39% RSD, respectively. Based on the results, the bioavailability of tungsten metal would be expected to be low for the oral, dermal, and inhalation routes of administration.

Executive summary:

Bioaccumulation potential cannot be judged based on study results.

Reference 7

Endpoint:

basic toxicokinetics in vitro / ex vivo

Type of information:

experimental study

Adequacy of study:

key study

Study period:

2009-01-28 To: 2009-05-01

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

test procedure in accordance with generally accepted scientific standards and described in sufficient detail

Remarks:

The bioaccessability study described in this report was performed according to the "Draft Guidance for RIP 3.6: Bioavailability and Read-Across for Metals and Minerals". GLP study.

Reason / purpose for cross-reference:

reference to other study

Objective of study:

absorption

Qualifier:

according to guideline

Guideline:

other: Draft Guidance for RIP 3.6: Bioavailability and Read-Across for Metals and Minerals

Deviations:

no

GLP compliance:

yes

Radiolabelling:

no

Species:

other: Human simulated fluids

Strain:

not specified

Sex:

not specified

Route of administration:

other: In vitro study

Details on exposure:

Preparation of Testing Fluids: The simulated fluids used in this study were gastric, lung alveolar, lung interstitial, lysosomal and artificial sweat. All salts and reagents used for the simulated fluid preparation were obtained from Sigma-Aldrich (St. Louis, MO).

Extraction Experiments: Extractions of the test substances in the simulated fluids were performed at pre-set time periods (up to 72 hours) while protected from light, using 0.1 g of sample in 50 ml of simulated fluid, at 37 °C and with continuous shaking (for all fluids except sweat, for which only initial shaking was performed). The experiments were carried out as follows:
- Simulated Gastric Fluid: The reactions were sampled for the determination of tungsten at 5 hours.
- Simulated Interstitial, Alveolar and Lysosomal Fluids: 5% CO2 in nitrogen was bubbled into solution at a rate of 50 ml/min. The reactions were sampled for the determination of tungsten at 2, 5, 24 and 72 hours.
- Simulated Sweat: The reactions were sampled for the determination of tungsten after 12 hours. No shaking was performed after the initial set up.

Sample Preparation: The simulated fluid extract samples were filtered immediately after sampling using 50 ml centrifuge tubes equipped with 0.45 microns PVDF filters (Grace Alltech, Deerfield, IL). The filtrates were stored in plastic bottles at 35 °C until analysis.

The extracts were then analysed for tungsten by inductively coupled plasma-mass spectrometry (ICP-MS).

Duration and frequency of treatment / exposure:

Single application of tungsten with fluids. Simulated Gastric Fluid was sampled for the determination of tungsten at 5 hours. Simulated Interstitial, Alveolar and Lysosomal Fluids were sampled for the determination of tungsten at 2, 5, 24 and 72 hours. Simulated Sweat was sampled for the determination of tungsten after 12 hours.

Remarks:

Doses / Concentrations:
0.1 g of test substance in 50 mL of simulated fluid

Control animals:

no

Details on dosing and sampling:

PHARMACOKINETIC STUDY (Absorption, distribution, excretion)
- Tissues and body fluids sampled: Simulated gastric fluid, Simulated interstitial fluid, Simulated alveolar fluid, Simulated lysosomal fluid and Simulated sweat
- Time and frequency of sampling: Simulated gastric fluid sampled at 5 hours, Simulated interstitial fluid sampled at 2, 5, 24 and 72 hours, Simulated alveolar fluid sampled at 2, 5, 24 and 72 hours, Simulated lysosomal fluid sampled at 2, 5, 24 and 72 hours and Simulated sweat sampled after 12 hours.

Sample analysis: Samples were diluted (if necessary), spiked with an internal standard [bismuth (Bi) at 1,000 pg/mL, prepared from dilution of a 1,000 ug/mL Certified Standard; Ultra Scientific, North Kingston, RI and analyzed directly on a Perkin Elmer Elan DRC II ICP-MS equipped with a dynamic reaction cell (DRC) and PerkinElmer AS-93 Plus autosamples instrument, according to methods established at IITRI for this study. A standard curve (prepared from dilutions of a 10,000 5%HNO3/6% HF; inorganic Ventures, Lakewood, NJ] was analyzed along with samples on each day of analysis. Instruments calibrators were prepared by diluting Certified Standard with 0.5% nitric acid to concentrations of approximately 200; 400; 800; 1,600; 3,200; 6,400; 13,000 and 25,000 pg/mL.

Statistics:

Calibration curves, regression coefficients and r-squared values were calculated using PerkinElmer ICP-MS software and Microsoft Excel software. Concentration values of tungsten in the study samples were calculated from linear regression coefficients derived from calibration standards that bracketed the expected concentration levels of tungsten in the study samples.

Details on absorption:

The fluid extracts were diluted 1:2500 for analysis. The average amount of tungsten found in the extracts (expressed as percentage of the test material) was in the 0.054 to 0.97% range. Percent relative standard deviations (%RSD; across all extraction times) ranged from 5.8 to 56%. Sweat Fluid: The percent of available tungsten in simulated sweat fluid sampled at 12 hours was 0.82 +/-0.14 % (18 % relative standard deviation).

Toxicokinetic parameters:

other: not applicable as this was an in vitro study

Metabolites identified:

not measured

Conclusions:

Interpretation of results (migrated information): bioaccumulation potential cannot be judged based on study results
The average amount of tungsten found in the extracts (expressed as percentage of the test material) was in the 0.054 to 0.97% range. Percent relative standard deviations (%RSD; across all extraction times) ranged from 5.8 to 56%. In the simulated sweat fluid, the percent of available tungsten sampled at 12 hours was 0.82 +/-0.14 % (18 % relative standard deviation). Based on the results, the bioavailability of tungsten metal would be expected to be low for the oral, dermal, and inhalation routes of administration.

Executive summary:

Bioaccumulation potential cannot be judged based on study results.

Reference 8

Endpoint:

basic toxicokinetics in vivo

Type of information:

experimental study

Adequacy of study:

supporting study

Reliability:

4 (not assignable)

Rationale for reliability incl. deficiencies:

documentation insufficient for assessment

Remarks:

Insufficient information provided on methods and results to accurately evaluate the study; no guideline available.

Reason / purpose for cross-reference:

reference to other study

Objective of study:

distribution

Qualifier:

no guideline followed

Principles of method if other than guideline:

Determination of the distribution of tungsten in various organs of the rat following oral ingestion of tungsten compounds.

GLP compliance:

no

Radiolabelling:

no

Species:

rat

Strain:

not specified

Sex:

male/female

Details on test animals or test system and environmental conditions:

Thirty-seven day old rats were placed on a basal diet of ground purina dog chow in which the finely-ground tungsten compounds had been incorporated. For each experimental diet, 2 female and 2 male rats/dose were used with the sexes separately caged. Diet and water were given ad libitum.

Route of administration:

oral: feed

Vehicle:

unchanged (no vehicle)

Details on exposure:

For each experimental diet, 2 female and 2 male rats/dose were used.

Duration and frequency of treatment / exposure:

Approximately one hundred days

Remarks:

Doses / Concentrations:
Diet E: 2.0% tungsten metal (2.0% W) in Purina dog chow
Diet F: 10. 0% tungsten metal (2.0% W) in Purina dog chow
Diet G: 10. 0% tungsten metal which had been further purified by multiple washes with 0.1N HCl and 0.1N NaOH followed by drying (2.0% W) in Purina dog chow

No. of animals per sex per dose / concentration:

2 female and 2 male

Control animals:

yes, concurrent no treatment

Positive control reference chemical:

no

Details on study design:

The tissues of both rats per sex on the same diet were combined, weighed, rinsed free of blood, and prepared for analysis. The method of Aull and Kinard was used for the determination of the tungsten.

Details on dosing and sampling:

PHARMACOKINETIC STUDY (Absorption, distribution, excretion)
- Tissues and body fluids sampled: brain, heart, uterus, lung, muscle, testis, blood, kidney, and liver

Statistics:

no data

Details on distribution in tissues:

Diet E: 2.0 % tungsten metal (2.0% W) in Purina dog chow -
No traces of tungsten were found in the brain, heart, or uterus. No relation was observed between diet or sex and the presence or absence of tungsten in the lungs, muscle, or testis. Trace amounts of tungsten were found in the blood, kidney and liver of both male and female rats. Ten and 10 mg p tungsten was found in the bone of male and female rats respectively. Two and 6 mg weight tungsten was found in the skin of male and female rats respectively. Trace amounts of tungsten were found in the male spleen and 2 mg where found in the female spleen.

Diet F: 10.0% tungsten metal (2.0% W) in Purina dog chow -
No traces of tungsten were found in the brain, heart, or uterus. No relation was observed between diet or sex and the presence or absence of tungsten in the lungs, muscle, or testis. Trace amounts of tungsten were found in the blood, kidney, and liver of both male and female rats. Fourteen and 18 mg tungsten was found in the bone of male and female rats respectively. Seven and 14 mg weight tungsten was found in the skin of male and female rats respectively. Trace amounts of tungsten were found in the male and female spleen.

Diet G: 2.0% tungsten metal (2.0% W) in Purina dog chow -
No traces of tungsten were found in the brain, heart of males, or uterus. A trace of tungsten was found in the heart of females. No relation was observed between diet or sex and the presence or absence of tungsten in the lungs, muscle, or testis. Trace amounts of tungsten were found in the blood, kidney and liver of both male and female rats. Eleven and 18 mg tungsten was found in the bone of male and female rats respectively. Eight and 9 mg weight tungsten was found in the skin of male and female rats respectively. Two mg and 3 mg tungsten were found in the male and female spleen, respectively.

Metabolites identified:

not measured

Description of key information

Limited in vivo toxicokinetic data were available on tungsten metal with the data that was available indicating the low bioavailability of tungsten when administered via the oral route. In a GLP in vitro study conducted according to Draft Guidance for RIP 3.6: Bioavailability and Read-Across for Metals and Minerals (Eurometaux 2008), the bioavailability of tungsten metal was determined in simulated human gastric, sweat, alveolar, lysosomal, and interstitial fluids. The average amount of tungsten found in the extracts (expressed as percentage of the test material) was in the 0.0546 to 0.971% range. The maximum solubility was determined at 72 hours for the simulated alveolar and lysosomal fluids (0.97%). Percent relative standard deviations (%RSD; across all extraction times) ranged from 5.86 to 56%. In the simulated gastric fluid, the percent of available tungsten sampled at 5 hours was 0.235 +/- 0.0135 % (5.8 % relative standard deviation). In the simulated sweat fluid, the percent of available tungsten sampled at 12 hours was 0.8290 +/-0.146 % (18 % relative standard deviation). In the simulated alveolar fluid, the percent of available tungsten sampled at 2, 5, 24, and 72 hours was 0.05460 +/- 0.00448%, 0.07280 +/- 0.00617%, 0.168 +/- 0.0134%, and 0.97+/- 0.134 %, respectively, with percent relative standard deviations of 8.0, 8.4, 8.0, and 13 %RSD, respectively. In the simulated lysosomal fluid, the percent of available tungsten sampled at 2, 5, 24, and 72 hours was 0.202 +/- 0.07280%, 0.325 +/- 0.168%, 0.4853 +/- 0.1820%, and 0.97+/- 0.3841 %, respectively, with percent relative standard deviations of 36, 52, 38, and 39% RSD, respectively. In the simulated interstitial fluid, the percent of available tungsten sampled at 2, 5, 24, and 72 hours was 0.07987 +/- 0.0168%, 0.0909 +/- 0.0325%, 0.179 +/- 0.09611%, and 0.719 +/- 0.3741 %, respectively, with percent relative standard deviations of 20, 36, 56, and 53% RSD, respectively. Based on the results, the bioavailability of tungsten metal would be expected to be low for the oral, dermal, and inhalation routes of administration.

 

In addition a toxicokinetic study conducted according to OECD 417 on tungsten oxide was used for read across. When tungsten oxide was administered via the inhalation route, tungsten was absorbed systemically with blood tungsten concentrations reaching maximum at 0 hours post-exposure. Cmax and AUC increased proportionally with increasing tungsten oxide dose level: Cmax was 0.819 ± 0.215 and 10.9 ± 4.7 fig/g, and AUC was 11.8 ± 3.29 and 148 ± 33.7 hr*Rg/g for the 0.08 and 0.65 mg/L dose groups, respectively. The systemic elimination half-life was 23 ± 4.3 and 154 ± 92.8 hours for the 0.08 and 0.65 mg/L dose groups, respectively, and the clearance rate was 1.24 ± 0.39 and 0.78 ± 0.18 L/hr/kg for the 0.08 and 0.65 mg/L dose groups, respectively. The majority of the inhaled tungsten oxide was excreted through the gastrointestinal tract. Tungsten concentration in lung tissue was at least one order of magnitude greater than in any other organ collected. In general, the femur and kidney ranked second and third highest, respectively, in tungsten concentration on a per organ weight basis. In terms of total tungsten burden in the organs, the lung, femur and liver ranked first, second and third, respectively. Tungsten distribution in tissue and organs reached steady-state on study Day 14. With the exception of the femur, the functional elimination half-life of tungsten in most organs was less than 24 hours

Key value for chemical safety assessment

Bioaccumulation potential:

no bioaccumulation potential

Additional information

No inhalation toxicokinetic data of sufficient quality are available for tungsten metal (target substance). However, toxicokinetic data are available for tungsten oxide (source substance), which are 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 in the Category section of this IUCLID submission or Annex 3 in the CSR.