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Endpoint:
basic toxicokinetics
Type of information:
experimental study
Adequacy of study:
supporting study
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
comparable to guideline study with acceptable restrictions
Remarks:
Similar to OECD Test Guideline 417 Toxicokinetics-distribution with acceptable deviations.
Objective of study:
distribution
Qualifier:
equivalent or similar to guideline
Guideline:
OECD Guideline 417 (Toxicokinetics)
Version / remarks:
: distribution
GLP compliance:
no
Radiolabelling:
no
Species:
mouse
Strain:
other: C57BL/6N
Sex:
female
Details on test animals or test system and environmental conditions:
- Body weight: 18 - 23 g
- Supplier: Hilltop Labs
Route of administration:
oral: gavage
Vehicle:
other: purified water
Details on exposure:
PREPARATION OF DOSING SOLUTIONS:
- Oral dose formulations were prepared on the day of administration by dilution of sodium tungstate dihydrate in purified water; these doses were selected to provide sufficient dose separation for the dose-response analysis of the disposition after oral administration.
- The doses span two log dilutions, and are all well within the tolerable concentration limits for W in rodents.
Duration and frequency of treatment / exposure:
single gavage administration
Remarks:
Doses / Concentrations:
- Oral doses were prepared and administered at 1, 10, or 100 mg/kg.
No. of animals per sex per dose / concentration:
4 female animals per dose.
Control animals:
yes
Details on study design:
Four female mice were exposed per dose group. One animal per dose group was then sacrificed at each time point. Animals were sacrificed at 1, 2, 4, and 24 hours.
Details on dosing and sampling:
COLLECTION OF SAMPLES:
- Animals were euthanized by an overdose of sodium pentobarbital administered by intraperitoneal injection.
- Plasma and tissues (intestine, liver, kidneys, femur, and uterus) were collected immediately after euthanasia (1, 2, 4, and 24 hrs after dosing).
- Plasma: blood was collected via cardiac puncture, placed in heparinized containers, and centrifuged at 2000 x g for 20 min.
- Tissues: liver, uterus, intestine (all three sections including contents), femur, and kidneys were harvested, weighed, placed in polypropylene containers, and stored at -70°C.

INSTRUMENTAL ANALYSIS:
- Tungsten concentrations were determined with the use of low-resolution ICP/MS following U.S. Environmental Protection Agency method 200.8 (U.S. EPA, 1994).
- When tissues and plasma were analyzed, the instrument response was recorded in micrograms of W per sample (ug/ sample).
- The data were then divided by the tissue weight to give micrograms of W per gram of tissue (ug/g), which is how the data in the current study are reported.
- In the case of plasma, direct weights were not recorded.
- Whole blood weights were recorded, and the amount of plasma was conservatively assigned a value of 55% of the whole blood weight.
Details on absorption:
W was readily absorbed from the gastrointestinal tract and distributed to each of the tissues analyzed.
Details on distribution in tissues:
- As shown by comparisons of the 100 mg/kg and 10 mg/kg and the 10 mg/kg and 1 mg/kg oral doses, W concentrations were not dose proportional in all biological matrices.
- The maximum concentrations were observed at 1-2 h for plasma and tissues.
- In all dose groups and time points the femur showed retention above background
- At 24 hours, W concentrations in most tissues for the low dose group were very close or at baseline levels. W concentrations for the mid and high doses were still above baseline levels, with the exception of the liver at both doses, the uterus at the high dose, and the intestine at mid dose.

No clinical abnormalities or gross tissue pathology were observed during tissue harvest from any of the studies.

Conclusions:
When administered orally, sodium tungstate as W was readily absorbed from the gastrointestinal tract and distributed to each of the tissues analyzed. The maximum concentrations were observed at 1-2 h for plasma and tissues. In all dose groups and time points the femur showed retention above background. At 24 hours, W concentrations in most tissues for the low dose group were very close or at baseline levels. W concentrations for the mid and high doses were still above baseline levels, with the exception of the liver at both doses, the uterus at the high dose, and the intestine at mid dose.
Endpoint:
basic toxicokinetics
Type of information:
experimental study
Adequacy of study:
supporting study
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
comparable to guideline study with acceptable restrictions
Remarks:
imilar to OECD Test Guideline 417 Toxicokinetics-distribution with acceptable deviations.
Objective of study:
distribution
Qualifier:
equivalent or similar to guideline
Guideline:
OECD Guideline 417 (Toxicokinetics)
Version / remarks:
: distribution
GLP compliance:
no
Radiolabelling:
no
Species:
rat
Strain:
Sprague-Dawley
Sex:
female
Details on test animals or test system and environmental conditions:
- Body weight: 240 - 265 g
- Supplier: Hilltop Labs
Route of administration:
oral: gavage
Vehicle:
other: purified water
Details on exposure:
PREPARATION OF DOSING SOLUTIONS:
- Oral dose formulations were prepared on the day of administration by dilution of sodium tungstate dihydrate in purified water; these doses were selected to provide sufficient dose separation for the dose-response analysis of the disposition after oral administration.
- The doses span two log dilutions, and are all well within the tolerable concentration limits for W in rodents.
Duration and frequency of treatment / exposure:
single gavage administration
Remarks:
Doses / Concentrations:
- Oral doses were prepared and administered at 1, 10, or 100 mg/kg.
No. of animals per sex per dose / concentration:
Four female rats were exposed per dose group. One animal per dose group was then sacrificed at each time point. Animals were sacrificed at 1, 2, 4, and 24 hours.
Control animals:
yes
Details on dosing and sampling:
COLLECTION OF SAMPLES:
- Animals were euthanized by an overdose of sodium pentobarbital administered by intraperitoneal injection.
- Plasma and tissues (intestine, liver, kidneys, femur, and uterus) were collected immediately after euthanasia (1, 2, 4, and 24 h after dosing).
- Plasma: blood was collected via cardiac puncture, placed in heparinized containers, and centrifuged at 2000 x g for 20 min.
- Tissues: liver, uterus, intestine (all three sections including contents), femur, and kidneys were harvested, weighed, placed in polypropylene containers, and stored at -70°C.

INSTRUMENTAL ANALYSIS:
- Tungsten concentrations were determined with the use of low-resolution ICP/MS following U.S. Environmental Protection Agency method 200.8
- When tissues and plasma were analyzed, the instrument response was recorded in micrograms of W per sample (ug/ sample).
- The data were then divided by the tissue weight to give micrograms of W per gram of tissue (ug/g), which is how the data in the current study are reported.
- In the case of plasma, direct weights were not recorded.
- Whole blood weights were recorded, and the amount of plasma was conservatively assigned a value of 55% of the whole blood weight.
Details on distribution in tissues:
- As shown by comparisons of 100 mg/kg and 10 mg/kg oral doses and the 10 mg/kg and 1 mg/kg oral doses, W concentrations increased with dose but were not always proportional to dose.
- At 24 h, W was not completely eliminated at the mid and high doses but returned to baseline levels at the low dose.
- A statistically significant difference in the W concentrations in plasma compared with whole blood was not observed, indicating that the plasma profile is a good indicator of blood concentrations and that W partitions equally between red blood cells and plasma.
-the W concentration in each of the rat tissues, with few exceptions, increases at each time point to a maximum at 4 h before becoming greatly reduced at 24 h.
- Intestine and kidneys at the high dose and femur at the mid-dose have a lower W concentration at 2 than at 1 h, but the concentrations still rose at 4 h before decreasing at 24 h.
- The other exceptions are intestine at the low dose, which had the highest W concentration at 2 h, and liver at the low dose, which had a steadily decreasing concentration

No clinical abnormalities or gross tissue pathology were observed during tissue harvest from any of the studies.

Conclusions:
As shown by comparisons of 100 mg/kg and 10 mg/kg oral doses and the 10 mg/kg and 1 mg/kg oral doses, W concentrations increased with dose but were not always proportional to dose. At 24 h, W was not completely eliminated at the mid and high doses but returned to baseline levels at the low dose. A statistically significant difference in the W concentrations in plasma compared with whole blood was not observed, indicating that the plasma profile is a good indicator of blood concentrations and that W partitions equally between red blood cells and plasma. The W concentration in each of the rat tissues, with few exceptions, increases at each time point to a maximum at 4 h before becoming greatly reduced at 24 h. Intestine and kidneys at the high dose and femur at the mid-dose have a lower W concentration at 2 than at 1 h, but the concentrations still rose at 4 h before decreasing at 24 h. The other exceptions are intestine at the low dose, which had the highest W concentration at 2 h, and liver at the low dose, which had a steadily decreasing concentration
Endpoint:
basic toxicokinetics
Type of information:
experimental study
Adequacy of study:
supporting study
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
comparable to guideline study with acceptable restrictions
Remarks:
Similar to OECD Test Guideline 417 Toxicokinetics-distribution with acceptable deviations.
Objective of study:
distribution
Qualifier:
equivalent or similar to guideline
Guideline:
OECD Guideline 417 (Toxicokinetics)
Version / remarks:
: distribution
Deviations:
yes
Remarks:
: only one dose group used
GLP compliance:
no
Radiolabelling:
no
Species:
mouse
Strain:
other: C57BL/6N
Sex:
female
Details on test animals or test system and environmental conditions:
- Body weight: 18 - 23 g
- Supplier: Hilltop Labs
Route of administration:
intravenous
Vehicle:
other: purified water
Details on exposure:
PREPARATION OF DOSING SOLUTIONS:
- Intravenous dose formulations were prepared on the day of administration by dilution of sodium tungstate dihydrate in purified water; these doses were selected to provide sufficient dose separation for the dose-response analysis of the disposition after oral administration.
- The doses span two log dilutions, and are all well within the tolerable concentration limits for W in rodents.
- Intravenous doses were prepared and at 1 mg/kg; this dose was selected to match the low oral dose.
Duration and frequency of treatment / exposure:
single intravenous dose
Remarks:
Doses / Concentrations:
- Intravenous doses were prepared and administered at 1 mg/kg.
No. of animals per sex per dose / concentration:
16 female animals total were administered the single dose. Four animals per time point were sacrificed.
Control animals:
yes
Details on dosing and sampling:
COLLECTION OF SAMPLES:
- Animals were euthanized by an overdose of sodium pentobarbital administered by intraperitoneal injection.
- Plasma and tissues (intestine, liver, kidneys, femur, and uterus) were collected immediately after euthanasia (1, 2, 4, and 24 h after dosing).
- Plasma: blood was collected via cardiac puncture, placed in heparinized containers, and centrifuged at 2000 x g for 20 min.
- Tissues: liver, uterus, intestine (all three sections including contents), femur, and kidneys were harvested, weighed, placed in polypropylene containers, and stored at -70°C.

INSTRUMENTAL ANALYSIS:
- Tungsten concentrations were determined with the use of low-resolution ICPMS following U.S. Environmental Protection Agency method 200.8
- When tissues and plasma were analyzed, the instrument response was recorded in micrograms of W per sample (µg/ sample).
- The data were then divided by the tissue weight to give micrograms of W per gram of tissue (µg/g), which is how the data in the current study are reported.
- In the case of plasma, direct weights were not recorded.
- Whole blood weights were recorded, and the amount of plasma was conservatively assigned a value of 55% of the whole blood weight.
Details on distribution in tissues:
- The liver, kidneys, and femur had a high concentration at 1 hour followed by a steady decrease through 24 hours.
- At 24 hours most of the tissues were very close to or at baseline W levels

- No clinical abnormalities or gross tissue pathology were observed during tissue harvest from any of the studies.

Conclusions:
The liver, kidneys, and femur had a high concentration at 1 hour followed by a steady decrease through 24 hours. At 24 hours most of the tissues were very close to or at baseline W levels
Endpoint:
basic toxicokinetics
Type of information:
experimental study
Adequacy of study:
key study
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
comparable to guideline study with acceptable restrictions
Remarks:
Similar to OECD Test Guideline 417 for toxicokinetics-distribution studies with acceptable deviations.
Reason / purpose for cross-reference:
reference to same study
Objective of study:
distribution
other: retention
Qualifier:
equivalent or similar to guideline
Guideline:
OECD Guideline 417 (Toxicokinetics)
Version / remarks:
: Section on Distribution Studies
GLP compliance:
no
Radiolabelling:
yes
Remarks:
[185W] tungstate
Species:
mouse
Strain:
NMRI
Sex:
female
Details on test animals or test system and environmental conditions:
- Virgin albino mice
- Feed: standard lab chow (Astra-Ewos, Sweden), available ad libitum
- Water: tap water, available ad libitum
- Temperature: 22 deg C
- Photoperiod: between 7 and 19 hours per day
Route of administration:
other: injection
Vehicle:
other: phosphate buffer
Details on exposure:
Preparation of dosing solutions:
- Two separate tungsten solutions were prepared for the injections of the mice.
- The isotope solution was diluted with phosphate buffer and the pH was adjusted with 0.2 M HCl to 7.2, so that each injection volume (about 0.1 mL) should contain 120 ug of [185W]/kg bw.
- The concentration of the isotope per injection volume was the same as for the first solution, but cold Na2WO4 was added to the phosphate buffer, so that each mouse would be exposed to about 20 mg of W/kg bw (= high dose). This tungsten concentration was similar to a concentration found in a previous study that produced fetal resorptions (Wide, 1984).

Mating procedure:
- Virgin females were caged with males overnight, and the day when a vaginal plug was found was taken as day 1 of gestation.

Duration and frequency of treatment / exposure:
Single injections.
Remarks:
Doses / Concentrations:
High dose = 20 mg tungsten/kg-body weight
Low dose = 120 ug tungsten/kg-body weight (for a mouse weighing 25 grams this corresponds to 27uCi)
Details on study design:
Autoradiography Experiments:
- On predetermined pregnancy days, corresponding to early organogenesis (day 8), mid-gestation (day 12) and late gestation (day 17), isotope injections were made in the tail vein. The animals were killed at various time intervals after the injections.
- Three mice were injected on day 8 of gestation with the low dose and killed after 4, 24, or 48 hours.
- One mouse received the high dose and was allowed to survive for 48 hours.
- Five mice were injected on gestation day 12, four with the low and one with the high dose; the low dose group was killed after 4, 24, or 48 hrs or 6 days (= day 18 of gestation), and the mouse given the high dose after 48 hrs.
- Two mice were given the low dose on day 17 of gestation and killed 4 or 24 hrs after.
- In addition, three male NMRI mice were taken to whole-body autoradiography after injection with the low dose; the males were killed at 1 hr, 24 hrs, or 10 days after the injection.
- The mice were killed by gaseous CO2 and if pregnant on day 17 or later, they were directly embedded in carboxymethylcellulose (CHIC) gel on a microtome stage and frozen by immersion in hexane cooled with dry ice.
- In the groups injected at early organogenesis or mid-gestation, the whole uteri (with ovaries) were excised, embedded, and frozen as above.
- The whole bodies or uteri were freeze-sectioned (20- and 60-µm-thick sections attached on tape), freeze-dried, and apposed against X-ray films (Industrex C, Kodak) for 4 to 16 weeks.

Quantitative Experiments:
- Mice on day 12 and 17 of gestation were administered iv [185W] tungstate at a dose level of 120 µg W/kg bw. They were then killed at 1 and 4 hrs (days 12 and 17) and at 48 hrs (day 12) after injection (four animals at each gestation stage and survival interval).
- In addition, eight mice at day 12 of gestation were administered iv a high dose (20 mg W/kg bw) together with the radioactive W. These animals were killed at 1 and 8 hrs after injection, four at each time point.
- At autopsy, serum and amniotic fluid plus a number of maternal organs, placentas, and whole fetuses were collected.
- The tissue samples were then dissolved in Soluene 350 (Packard) and scintillation fluid (4.9 mg PPO and 0.1 mg POPOP/liter toluene) was added for subsequent counting in a Packard Tri-Carb Model 2405.
- Serum and amniotic fluid samples were added to 1 mL of water and 10 mL of xnstagel (Packard); quenching was corrected for by the use of an external standard.
Statistics:
- The mean fetal and placental concentrations within each mother were first determined; these values were used as the sampling units in the statistic evaluation (Student's t test).
Details on distribution in tissues:
Adults:
- The distribution pattern of [185] W given in tracer doses to adult male as well as a pregnant female mice, was characterized by an accumulation of radioactivity in the skeleton, kidney, and liver, with rapid excretion to the urine and intestinal contents.
- Intestinal contents were high despite the fact that the concentration in bile was low, even compared to blood.
- Relatively high concentrations were found in the thyroid, adrenal medulla, and outer zone of the adrenal cortex, and pituitary; high activity was found also in cartilage.
- In the male, considerable accumulation occurred in the seminal vesicles.
- In the female, the interstitial tissues and-at long survival intervals-the follicles of the ovaries showed relatively high concentrations of tungsten.
- With time, radioactivity was cleared from the body with retention of activity, particularly in the skeleton, renal cortex, red pulp of the spleen, seminal vesicles of males, and previously mentioned organs such as the adrenal medulla and thyroid.
- There was a retention as well in the esophageal part of the stomach.
- In a pigmented mouse (C57BL), an accumulation of radioactivity occurred in the ciliary body, iris, and along the retina, which was not observed in albino mice; indicating an accumulation in melanin-containing structures.

Fetoplacental Unit:
- Day-8 treated group: Accumulation of [185]W was detected in the zone of the ectoplacental cone (the developing chorioallantoic placenta), in the visceral yolk sac epithelium, and in the decidua basalis at the 24 hr-survival interval
- Embryonic uptake was low; however, it was difficult to evaluate due to the small size of the embryo. The distribution was much the same after 48 hrs, the highest concentration found in the decidua basalis

Day-12 injected group: At this stage of gestation, the embryonic uptake was considerably higher than at Day 8, as judged by autoradiography. The fetal concentration was higher at 1 hr after injection than at 4 hrs. At 4 hrs after injection (the earliest time studied by autoradiography), there was an accumulation in the nervous tissues of the fetuses, while the activity in the rest of the body was evenly distributed. A high concentration was noted also in the visceral yolk sac epithelium. After 24 and 48 hrs, labeling of the fetal tissue had decreased, while some of the isotope was retained in the visceral yolk sac epithelium. The activity in the amniotic fluid was remarkably high, compared to serum, from 4 to 48 hr after injection. A retention of the isotope could still be observed in the visceral yolk sac and faintly in the fetal skeleton 5 days after the injection.
Day-17 injected group: The average fetal concentration was higher at Day 17 than at Day 12 of gestation, both at 1 and at 4 hrs after injection. As was the case at day 12 and also at Day 17, a rapid fall in fetal concentration occurred between 1 and 4 hrs postinjection. The fetal distribution pattern was dominated by an uptake in the skeleton at 4 hrs and even more at 24 hrs after injection. The concentration in fetal brain was low (cf. Day-12 injected mice). A notable uptake in fetal kidney was observed. As for Day 12 and also at Day 17, an accumulation was found in the amniotic fluid.
- High-dose experiments: High doses of W given to the pregnant mice resulted in lower relative concentrations (% of dose) in maternal organs (liver and kidney), while those of the fetuses, placentas, and amniotic fluids, as well as maternal serum and brain, were equivalent to those of the low-dose group at 1 hr after injection. At 4 hrs there was an even more pronounced decrease in the relative concentrations of maternal organs. At this time, even the amniotic fluid and-to some extent-even the placenta and fetus had lower relative concentrations as compared to the low-dose groups.
Conclusions:
Whole-body autoradiography and impulse counting experiments were used to study the distribution of radioactivity in the pregnant mouse after administration of [185] W tungstate. A rapid uptake was found in a number of tissues-skeleton, red pulp of the spleen, adrenal, liver, thyroid, pituitary, and ovary-and in the intestine and kidneys, through which it was rapidly excreted. [185] W was readily transported from mother to fetus, although more in late as compared to early gestation. The largest metal retention was found in the maternal skeleton, kidneys, spleen, and in the visceral yolk sac epithelium and the skeleton of the fetus.
Endpoint:
basic toxicokinetics
Type of information:
experimental study
Adequacy of study:
supporting study
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
comparable to guideline study with acceptable restrictions
Remarks:
Similar to OECD Test Guideline 417 Toxicokinetics-distribution with acceptable deviations.
Objective of study:
distribution
Qualifier:
equivalent or similar to guideline
Guideline:
OECD Guideline 417 (Toxicokinetics)
Version / remarks:
: distribution
Deviations:
yes
Remarks:
: only one dose used
GLP compliance:
no
Radiolabelling:
no
Species:
rat
Strain:
Sprague-Dawley
Sex:
female
Details on test animals or test system and environmental conditions:
- Body weight: 240 - 265 g
- Supplier: Hilltop Labs
Route of administration:
intravenous
Vehicle:
other: purified water
Details on exposure:
PREPARATION OF DOSING SOLUTIONS:
- Intravenous dose formulations were prepared on the day of administration by dilution of sodium tungstate dihydrate in purified water; these doses were selected to provide sufficient dose separation for the dose-response analysis of the disposition after oral administration.
- The doses span two log dilutions, and are all well within the tolerable concentration limits for W in rodents.
- Intravenous doses were prepared and at 1 mg/kg; this dose was selected to match the low oral dose.
Remarks:
Doses / Concentrations:
- Intravenous doses were prepared and administered at 1 mg/kg.
No. of animals per sex per dose / concentration:
16 female animals total. 4 animals were sacrificed at each time point.
Control animals:
yes
Details on dosing and sampling:
COLLECTION OF SAMPLES:
- Animals were euthanized by an overdose of sodium pentobarbital administered by intraperitoneal injection.
- Plasma and tissues (intestine, liver, kidneys, femur, and uterus) were collected immediately after euthanasia (1, 2, 4, and 24 h after dosing).
- Plasma: blood was collected via cardiac puncture, placed in heparinized containers, and centrifuged at 2000 x g for 20 min.
- Tissues: liver, uterus, intestine (all three sections including contents), femur, and kidneys were harvested, weighed, placed in polypropylene containers, and stored at -70°C.

INSTRUMENTAL ANALYSIS:
- Tungsten concentrations were determined with the use of low-resolution ICPMS following U.S. Environmental Protection Agency method 200.8
- When tissues and plasma were analyzed, the instrument response was recorded in micrograms of W per sample (ug/ sample).
- The data were then divided by the tissue weight to give micrograms of W per gram of tissue (ug/g), which is how the data in the current study are reported.
- In the case of plasma, direct weights were not recorded.
- Whole blood weights were recorded, and the amount of plasma was conservatively assigned a value of 55% of the whole blood weight.
Details on distribution in tissues:
- The highest W concentrations for each of the tissues, except intestine, were at 1 hour and steadily decreased through 24 hours.
- At 24 hours most of the rat tissue values were very close to or at baseline W levels.

- No clinical abnormalities or gross tissue pathology were observed during tissue harvest from any of the studies.

Conclusions:
The highest W concentrations for each of the tissues, except intestine, were at 1 hour and steadily decreased through 24 hours. At 24 hours most of the rat tissue values were very close to or at baseline W levels.
Endpoint:
basic toxicokinetics
Type of information:
experimental study
Adequacy of study:
supporting study
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
guideline study with acceptable restrictions
Remarks:
Well documented, scientifically sound study that follows OECD Guideline "417- Toxicokinetics", with a few deviations: housing conditions and acclimatization of animals were not discussed, and only distribution parameters were investigated (absorption, excretion, or metabolism were not examined).
Reason / purpose for cross-reference:
reference to same study
Objective of study:
distribution
Qualifier:
equivalent or similar to guideline
Guideline:
OECD Guideline 417 (Toxicokinetics)
Deviations:
yes
GLP compliance:
not specified
Radiolabelling:
no
Species:
mouse
Strain:
other: C57BL/6N
Sex:
female
Details on test animals or test system and environmental conditions:
TEST ANIMALS
- Source: Hilltop Lab Animals Inc. (Scottdale, PA)
- Age at study initiation: N/A
- Weight at study initiation: 17-20 g
- Fasting period before study: N/A
- Housing: N/A
- Individual metabolism cages: no data
- Diet (e.g. ad libitum): ad libitum
- Water (e.g. ad libitum): ad libitum
- Acclimation period: N/A


ENVIRONMENTAL CONDITIONS
- Temperature (°C):N/A
- Humidity (%): N/A
- Air changes (per hr): N/A
- Photoperiod (hrs dark / hrs light):N/A

Route of administration:
oral: drinking water
Details on exposure:
PREPARATION OF DOSING SOLUTIONS: Drinking water doses were prepared every third day at 560 mg/L and administered ad libitum. Water bottles containing 560 mg/L tungstate were weighed prior to placing in cages. After 3 days, the water bottles were weighed again and the difference was considered the volume of the solution consumed in that time period. The weight of administered tungsten was calculated and divided by the animal's body weight to give the administered dose.


Duration and frequency of treatment / exposure:
14 days, ad libitum
Remarks:
Doses / Concentrations:
560 mg/L (actual dose= 3951.8 +/- 216.7 mg/kg)
No. of animals per sex per dose / concentration:
4 animals
Control animals:
other: yes, (n=6) no specific data regarding control treatment
Details on study design:
- Dose selection rationale: Doses were selected to correspond with ongoing NTP studies.
Details on dosing and sampling:
PHARMACOKINETIC STUDY (distribution)
- Tissues and body fluids sampled: plasma, intestines, liver, kidneys, femur, uterus
- Time and frequency of sampling: tissues were harvested immediately after euthanasia, after the 14 days of exposure





Details on distribution in tissues:
There was a significant increase in tungsten tissue concentration for treated mice (vs. endogenous concentrations-control) in plasma and all tissues examined (intestine, liver, kidneys, femur, and uterus).
Conclusions:
After ad libitum drinking water exposure to 560 mg/L (actual dose= 3951.8 +/- 216.7 mg/kg) of the test substance for 14 days, female C57B1/6N mice (vs. endogenous concentrations-control) exhibited significantly elevated tungsten concentrations in plasma and all tissues examined (intestine, liver, kidneys, femur, and uterus).
Endpoint:
basic toxicokinetics
Type of information:
experimental study
Adequacy of study:
supporting study
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
guideline study with acceptable restrictions
Remarks:
Well documented, scientifically sound study that follows OECD Guideline "417- Toxicokinetics", with a few deviations: housing conditions and acclimatization of animals were not discussed, and only distribution parameters were investigated (absorption, excretion, or metabolism were not examined).
Reason / purpose for cross-reference:
reference to same study
Objective of study:
distribution
Qualifier:
equivalent or similar to guideline
Guideline:
OECD Guideline 417 (Toxicokinetics)
Deviations:
yes
GLP compliance:
not specified
Radiolabelling:
no
Species:
mouse
Strain:
other: C57BL/6N
Sex:
female
Details on test animals or test system and environmental conditions:
TEST ANIMALS
- Source: Hilltop Lab Animals Inc. (Scottdale, PA)
- Age at study initiation: N/A
- Weight at study initiation: 17-20 g
- Fasting period before study: N/A
- Housing: N/A
- Individual metabolism cages: no data
- Diet (e.g. ad libitum): ad libitum
- Water (e.g. ad libitum): ad libitum
- Acclimation period: N/A


ENVIRONMENTAL CONDITIONS
- Temperature (°C):N/A
- Humidity (%): N/A
- Air changes (per hr): N/A
- Photoperiod (hrs dark / hrs light):N/A

Route of administration:
oral: drinking water
Details on exposure:
PREPARATION OF DOSING SOLUTIONS: Drinking water doses were prepared every third day at 560 mg/L and administered ad libitum. Water bottles containing 560 mg/L tungstate were weighed prior to placing in cages. After 3 days, the water bottles were weighed again and the difference was considered the volume of the solution consumed in that time period. The weight of administered tungsten was calculated and divided by the animal's body weight to give the administered dose.

Gestating females were administered drinking water containing tungsten starting at day 6 of confirmed gestation, through the duration of the study, the days of primary organogenesis.
Duration and frequency of treatment / exposure:
9 days, ad libitum
Remarks:
Doses / Concentrations:
560 mg/L (actual dose= 1238.5 +/- 186.1 mg/kg)
No. of animals per sex per dose / concentration:
4
Control animals:
other: yes, (n=6) no specific data regarding control treatment
Details on study design:
- Dose selection rationale: Doses were selected to correspond with ongoing NTP studies.
Details on dosing and sampling:
PHARMACOKINETIC STUDY (distribution)
- Tissues and body fluids sampled: plasma, intestines, liver, kidneys, femur, uterus, fetus
- Time and frequency of sampling: tissues were harvested immediately after euthanasia, after the 9 days of exposure





Details on distribution in tissues:
There was a significant increase in tungsten tissue concentration for treated mice (vs. endogenous concentrations-control) in plasma and all tissues examined (intestine, liver, kidneys, femur, and uterus). Fetuses also showed a significantly increased tungsten concentration above endogenous levels.
Conclusions:
After ad libitum drinking water exposure to 560 mg/L (actual dose= 1238.5 +/- 186.1 mg/kg) of the test substance for 9 days, female C57B1/6N mice (vs. endogenous concentrations-control) exhibited significantly elevated tungsten concentrations in plasma and all tissues examined (intestine, liver, kidneys, femur, and uterus). In addition, fetuses also showed a significantly increased tungsten concentration above endogenous levels.
Endpoint:
basic toxicokinetics
Type of information:
experimental study
Adequacy of study:
supporting study
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
guideline study with acceptable restrictions
Remarks:
Follows OECD Guideline "417- Toxicokinetics", with a few deviations: housing conditions and acclimatization of animals were not discussed, and only distribution parameters were investigated (absorption, excretion, or metabolism were not examined). The study was not GLP certified.
Reason / purpose for cross-reference:
reference to same study
Objective of study:
distribution
Qualifier:
equivalent or similar to guideline
Guideline:
OECD Guideline 417 (Toxicokinetics)
Deviations:
yes
GLP compliance:
not specified
Radiolabelling:
no
Species:
rat
Strain:
Sprague-Dawley
Sex:
female
Details on test animals or test system and environmental conditions:
TEST ANIMALS
- Source: Hilltop Lab Animals Inc. (Scottdale, PA)
- Age at study initiation: N/A
- Weight at study initiation: 223-271 g
- Fasting period before study: N/A
- Housing: N/A
- Individual metabolism cages: no data
- Diet (e.g. ad libitum): ad libitum
- Water (e.g. ad libitum): ad libitum
- Acclimation period: N/A


ENVIRONMENTAL CONDITIONS
- Temperature (°C):N/A
- Humidity (%): N/A
- Air changes (per hr): N/A
- Photoperiod (hrs dark / hrs light):N/A

Route of administration:
oral: drinking water
Details on exposure:
PREPARATION OF DOSING SOLUTIONS: Drinking water doses were prepared every third day at 560 mg/L and administered ad libitum. Water bottles containing 560 mg/L tungstate were weighed prior to placing in cages. After 3 days, the water bottles were weighed again and the difference was considered the volume of the solution consumed in that time period. The weight of administered tungsten was calculated and divided by the animal's body weight to give the administered dose.

Duration and frequency of treatment / exposure:
14 days, ad libitum
Remarks:
Doses / Concentrations:
560 mg/L (actual dose= 1224.7 +/- 140.7 mg/kg)
No. of animals per sex per dose / concentration:
4
Control animals:
other: yes, (n=6) no specific data regarding control treatment
Details on study design:
- Dose selection rationale: Doses were selected to correspond with ongoing NTP studies.
Details on dosing and sampling:
PHARMACOKINETIC STUDY (distribution)
- Tissues and body fluids sampled: plasma, intestines, liver, kidneys, femur, uterus
- Time and frequency of sampling: tissues were harvested immediately after euthanasia, after the 14 days of exposure





Details on distribution in tissues:
There was a significant increase in tungsten tissue concentration for treated rats (vs. endogenous concentrations-control) in plasma and all tissues examined (intestine, liver, kidneys, femur, and uterus).
Conclusions:
After ad libitum drinking water exposure to 560 mg/L (actual dose= 1224.7 +/- 140.7 mg/kg) of the test substance for 14 days, female Sprague-Dawley rats (vs. endogenous concentrations-control) exhibited significantly elevated tungsten concentrations in plasma and all tissues examined (intestine, liver, kidneys, femur, and uterus).
Endpoint:
basic toxicokinetics
Type of information:
experimental study
Adequacy of study:
supporting study
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
guideline study with acceptable restrictions
Remarks:
Well documented, scientifically sound study that follows OECD Guideline "417- Toxicokinetics", with a few deviations: housing conditions and acclimatization of animals were not discussed, and only distribution parameters were investigated (absorption, excretion, or metabolism were not examined).
Reason / purpose for cross-reference:
reference to same study
Objective of study:
distribution
Qualifier:
equivalent or similar to guideline
Guideline:
OECD Guideline 417 (Toxicokinetics)
Deviations:
yes
GLP compliance:
not specified
Radiolabelling:
no
Species:
rat
Strain:
Sprague-Dawley
Sex:
female
Details on test animals or test system and environmental conditions:
TEST ANIMALS
- Source: Hilltop Lab Animals Inc. (Scottdale, PA)
- Age at study initiation: N/A
- Weight at study initiation: 223-271 g
- Fasting period before study: N/A
- Housing: N/A
- Individual metabolism cages: no data
- Diet (e.g. ad libitum): ad libitum
- Water (e.g. ad libitum): ad libitum
- Acclimation period: N/A


ENVIRONMENTAL CONDITIONS
- Temperature (°C):N/A
- Humidity (%): N/A
- Air changes (per hr): N/A
- Photoperiod (hrs dark / hrs light):N/A

Route of administration:
oral: drinking water
Details on exposure:
PREPARATION OF DOSING SOLUTIONS: Drinking water doses were prepared every third day at 560 mg/L and administered ad libitum. Water bottles containing 560 mg/L tungstate were weighed prior to placing in cages. After 3 days, the water bottles were weighed again and the difference was considered the volume of the solution consumed in that time period. The weight of administered tungsten was calculated and divided by the animal's body weight to give the administered dose.

Gestating females were administered drinking water containing tungsten starting at day 6 of confirmed gestation, through the duration of the study, the days of primary organogenesis.
Duration and frequency of treatment / exposure:
10 days, ad libitum
Remarks:
Doses / Concentrations:
560 mg/L (actual dose= 1067.9 +/- 299.7 mg/kg)
No. of animals per sex per dose / concentration:
4
Control animals:
other: yes, (n=6) no specific data regarding control treatment
Details on study design:
- Dose selection rationale: Doses were selected to correspond with ongoing NTP studies.
Details on dosing and sampling:
PHARMACOKINETIC STUDY (distribution)
- Tissues and body fluids sampled: plasma, intestines, liver, kidneys, femur, uterus, fetus
- Time and frequency of sampling: tissues were harvested immediately after euthanasia, after the 10 days of exposure





Details on distribution in tissues:
There was a significant increase in tungsten tissue concentration for treated rats (vs. endogenous concentrations-control) in plasma and all tissues examined (intestine, liver, kidneys, femur, and uterus). Fetuses also showed increased tungsten concentration above endogenous levels, though it was not indicated whether or not this increase was significant.
Conclusions:
After ad libitum drinking water exposure to 560 mg/L (actual dose was 1067.9 +/- 299.7 mg/kg) of the test substance for 10 days, female Sprague-dawley rats (vs. endogenous concentrations-control) exhibited significantly elevated tungsten concentrations in plasma and all tissues examined (intestine, liver, kidneys, femur, and uterus). In addition, fetuses also showed increased tungsten concentration above endogenous levels.
Endpoint:
basic toxicokinetics
Type of information:
experimental study
Adequacy of study:
supporting study
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
guideline study with acceptable restrictions
Remarks:
Well documented, scientifically sound study that follows OECD Guideline "417- Toxicokinetics", with a few deviations: housing conditions and acclimatization of animals were not discussed, and only distribution parameters were investigated (absorption, excretion, or metabolism were not examined).
Reason / purpose for cross-reference:
reference to same study
Objective of study:
distribution
Qualifier:
equivalent or similar to guideline
Guideline:
OECD Guideline 417 (Toxicokinetics)
Deviations:
yes
GLP compliance:
not specified
Radiolabelling:
no
Species:
mouse
Strain:
other: C57BL/6N
Sex:
female
Details on test animals or test system and environmental conditions:
TEST ANIMALS
- Source: Hilltop Lab Animals Inc. (Scottdale, PA)
- Age at study initiation: N/A
- Weight at study initiation: 17-20 g
- Fasting period before study: N/A
- Housing: N/A
- Individual metabolism cages: no data
- Diet (e.g. ad libitum): ad libitum
- Water (e.g. ad libitum): ad libitum
- Acclimation period: N/A


ENVIRONMENTAL CONDITIONS
- Temperature (°C):N/A
- Humidity (%): N/A
- Air changes (per hr): N/A
- Photoperiod (hrs dark / hrs light):N/A

Route of administration:
oral: gavage
Vehicle:
water
Details on exposure:
PREPARATION OF DOSING SOLUTIONS: Prepared on the day of administration by the dilution of sodium tungstate dihydrate in purified water.


Duration and frequency of treatment / exposure:
14 days, daily
Remarks:
Doses / Concentrations:
10 mg/kg (actual dose= 10.2 +/- 0.3 mg/kg)
No. of animals per sex per dose / concentration:
4
Control animals:
other: yes, (n=6) no specific data regarding control treatment
Details on study design:
- Dose selection rationale: Doses were selected to correspond with ongoing NTP studies.
Details on dosing and sampling:
PHARMACOKINETIC STUDY (distribution)
- Tissues and body fluids sampled: plasma, intestines, liver, kidneys, femur, uterus
- Time and frequency of sampling: tissues were harvested immediately after euthanasia, after the 14 days of dosing





Details on distribution in tissues:
There was a slight increase in tungsten tissue concentration for treated rodents (vs. endogenous concentrations-control) in the plasma, intestines, liver, and femur after 14 days. No difference in concentration was observed in kidneys. In contrast, the uterus displayed a large increase in tungsten concentration and was significantly above endogenous (control) levels.
Conclusions:
After oral exposure to 10mg/kg/day (actual dose= 10.2 +/- 0.3 mg/kg) of the test substance for 14 days, female C57B1/6N mice (vs. endogenous concentrations-control) exhibited significantly elevated tungsten concentrations in uterus tissue.
Endpoint:
basic toxicokinetics
Type of information:
experimental study
Adequacy of study:
supporting study
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
guideline study with acceptable restrictions
Remarks:
Well documented, scientifically sound study that follows OECD Guideline "417- Toxicokinetics", with a few deviations: housing conditions and acclimatization of animals were not discussed, and only distribution parameters were investigated (absorption, excretion, or metabolism were not examined).
Reason / purpose for cross-reference:
reference to same study
Objective of study:
distribution
Qualifier:
equivalent or similar to guideline
Guideline:
OECD Guideline 417 (Toxicokinetics)
Deviations:
yes
GLP compliance:
not specified
Radiolabelling:
no
Species:
rat
Strain:
Sprague-Dawley
Sex:
female
Details on test animals or test system and environmental conditions:
TEST ANIMALS
- Source: Hilltop Lab Animals Inc. (Scottdale, PA)
- Age at study initiation: N/A
- Weight at study initiation: 223-271g
- Fasting period before study: N/A
- Housing: N/A
- Individual metabolism cages: no data
- Diet (e.g. ad libitum): ad libitum
- Water (e.g. ad libitum): ad libitum
- Acclimation period: N/A


ENVIRONMENTAL CONDITIONS
- Temperature (°C):N/A
- Humidity (%): N/A
- Air changes (per hr): N/A
- Photoperiod (hrs dark / hrs light):N/A

Route of administration:
oral: gavage
Vehicle:
water
Details on exposure:
PREPARATION OF DOSING SOLUTIONS: Prepared on the day of administration by the dilution of sodium tungstate dihydrate in purified water.


Duration and frequency of treatment / exposure:
14 days, daily
Remarks:
Doses / Concentrations:
10 mg/kg (actual dose= 9.4 +/- 0.3 mg/kg)
No. of animals per sex per dose / concentration:
4
Control animals:
other: yes, (n=6) no specific data regarding control treatment
Details on study design:
- Dose selection rationale: Doses were selected to correspond with ongoing NTP studies.
Details on dosing and sampling:
PHARMACOKINETIC STUDY (distribution)
- Tissues and body fluids sampled: plasma, intestines, liver, kidneys, femur, uterus
- Time and frequency of sampling: tissues were harvested immediately after euthanasia, after the 14 days of dosing





Details on distribution in tissues:
There was an increase in tungsten tissue concentration for treated rodents (vs. endogenous concentrations-control) in plasma, and all tissues examined. The plasma, liver, and uterus showed tungsten levels only slighty above baseline levels; however, the intestine, kidneys, and femur displayed significant increases in the amount of tungsten.
Conclusions:
After oral exposure to 10mg/kg (actual dose= 9.4 +/- 0.3 mg/kg) of the test substance for 14 days, female Sprague-Dawley rats (vs. endogenous concentrations-control) exhibited significantly elevated tungsten concentrations in intestine, kidney, and femur tissue.
Endpoint:
basic toxicokinetics
Type of information:
experimental study
Adequacy of study:
key study
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
guideline study with acceptable restrictions
Remarks:
Well documented study with methods similar to OECD guideline 417.
Objective of study:
distribution
excretion
Qualifier:
equivalent or similar to guideline
Guideline:
OECD Guideline 417 (Toxicokinetics)
GLP compliance:
no
Radiolabelling:
yes
Remarks:
1.5-2.3 µCi of 181W as sodium tungstate
Species:
dog
Strain:
Beagle
Sex:
male/female
Route of administration:
intravenous
Vehicle:
unchanged (no vehicle)
Details on exposure:
Injections were made with a 1ml disposable syringe with a 25 gauge needle. Injected activities were determined by weighing the syringe before and after injection. The dogs were injected in the cephalic vein of the right foreleg.
Duration and frequency of treatment / exposure:
Two injections, one at the start of the study and the second 43 days after the first injection.
Remarks:
Doses / Concentrations:
Sodium tungstate injection solution was prepared from a solution of 181W (VI) in 0.5 N HCl plus 0.1 N HF with a specific activity of 792 µCi/ml which contained 0.4 mg tungsten/mL. 1.1944 g of this solution was added to 10 mg of non-radioactive tungstic acid dissolved in concentrated NH4OH. 5 ml of 30% H2O2 was added and the solution made normal by addition of HNO3. The solution was heated to 90°C. The dried tungstic acid was calcined at 850°C to produce 9.6 mg of tungstic oxide. This was fused with an excess of NaCO3 over a gas flame. The resulting sodium tungstate was dissolved in a small amount(~0.5 ml) of hot distilled water and made up to a volume of 10 ml with normal saline. The sodium tungstate solution was found to contain 26.8 µCi/g. This solution was further diluted with normal saline to give an injection solution containing 1.46±0.04 µCi/g.
No. of animals per sex per dose / concentration:
1 male and 1 female beagle
Control animals:
not specified
Details on excretion:
99% of the injected activity was excreted in the urine by 24 hours after injection.

Injected 181W sodium tungstate was removed from the body as if 81.6 % was cleared with an 86 min biological half-time, 15 % with an 8.8 hr half-time, 2 % with a 3.65 day biological half-time and 1 % with a 99 day biological half time. Loss of activity from the blood during the first 24 hours was very rapid. 70% of the activity in the blood was removed with a half-time of 36 minutes, 25% with a half-time of 71 minutes and 4.6% with a half-time of 5 hours.

Ninety-one percent of the injected activity was excreted in the urine by 24 hours after injection.

70% of the activity in the blood was removed with a half-time of 36 minutes, 25% with a half-time of 71 minutes, and 4.6% with a half-time of 5 hours. blood activity had declined to a value of less than the detection limit by 36 hours after detection. The average plasma plasma to red blood cell ration was 3:1 for the assay. There was no clear trend toward accumulation of activity associated with red blood cells.

Conclusions:
The IV infused radiolabelled sodium tungstate was rapidly removed from the blood with 36 hours to below detectable levels, with 99% of the injected activity excreted in the urine by 24 hours after injection.
Endpoint:
basic toxicokinetics in vivo
Type of information:
experimental study
Adequacy of study:
supporting study
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
guideline study with acceptable restrictions
Remarks:
Well documented study with sufficient information provided on methods and results to evaluate data.
Reason / purpose for cross-reference:
reference to same study
Objective of study:
distribution
excretion
Qualifier:
no guideline available
Principles of method if other than guideline:
The concentration of tungsten was measured in plasma samples after intravenous administration of sodium tungstate to male beagle dogs.
GLP compliance:
not specified
Radiolabelling:
no
Species:
dog
Strain:
Beagle
Sex:
male/female
Details on test animals or test system and environmental conditions:
TEST ANIMALS
- Source: Harlan (Grannat, France)
- Weight at study initiation: Mean weight 10.5 kg
- Fasting period before study: Yes
- Housing: housed individually during the study in stainless steel cages
- Diet: access to pelleted food (400 g/day per animal); Usine d’Alimentation Rationnelle, Villemoisson sur Orge, France)
- Water: Tap water was distributed ad libitum.
- Acclimation period: The animals underwent an acclimatization period of 10 days before the experiment.

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 18–21degree C
- Humidity (%): 40–70%
- Photoperiod (hrs dark / hrs light): constant 12 h light/dark cycle (lights on at 08.00 h)- artificial lighting
Route of administration:
intravenous
Vehicle:
other: 0.9% NaCl in aqueous solution
Details on exposure:
For intravenous administration, Sodium tungstate was administered to animals in an aqueous solution containing 0.9% sodium chloride. Six male dogs received two single doses of sodium tungstate (25 and 50 mg/kg) by intravenous route separated by at least a 15-day wash-out period (experiment 1). In intravenous exposures, sodium tungstate was administered over 1 minute into the cephalic vein.
Duration and frequency of treatment / exposure:
Two doses were administered over 1 min. into the cephalic vein.
Remarks:
Doses / Concentrations:
25 and 50 mg/kg
No. of animals per sex per dose / concentration:
six male dogs
Control animals:
no
Positive control reference chemical:
no
Details on study design:
During the treatment period, each dog was observed daily on at least two occasions and the fluid and food intake were measured. The animals were weighed twice a week in order to calculate daily doses. Laboratory studies including hematology, blood chemistry, and urinalysis were performed at baseline, after 6 weeks of treatment, and at the end of treatment.

Tungsten was quantified using an inductively coupled plasma emission spectrometric method at a wavelength of 207.91 nm. Calibration curves were obtained in the range 134–1300 ng/mL. Precision ranged from 0.4 to 17%, and accuracy was between 89 and 105%. Dilution has no influence on the performance of the method, which could then be used to quantify plasma containing up to 90 mg/mL. The limit of quantification was 100 ng/mL (precision, 17%). Plasma concentrations less than the limit of quantitation were omitted from the analysis.

Individual pharmacokinetic parameters were estimated using an empirical Bayes methodology. The population analysis was performed using the P-PHARM computer program (version 1.4, SIMED, Cre´teil, France). The population estimation algorithm is an EM-type procedure. The kinetic parameters considered in the population analysis consists of total clearance (CL), initial volume of distribution (V), transfer rate constants (k12 and k21 ), absorption rate constant (ka ), lag-time (tlag ) and bioavailability (F). Several secondary pharmacokinetic parameters were calculated from the individual (Bayesian estimates) primary pharmacokinetic parameters: the model-predicted maximum concentration (C ) and the time of peak (tmax), the area under the plasma concentration–time curve (AUC); the elimination half-life (t1/2elim ); and the of distribution (Vd-β). Moreover, the plasma concentration of tungsten at any time can be easily predicted using the empirical Bayes estimate of the pharmacokinetic parameters.

To determine model acceptance, predicted concentrations (Cpred ) for each individual were plotted versus observed concentrations (Cobs ). Residuals (Cpred –C obs) were plotted versus time and versus predicted concentrations. Then, a standardized concentration prediction error (SCPE) was calculated as follows: SCPE = [Cobs -Cpred]/SD(Cpred), where SD(Cpred ) represents the estimated standard deviation on the predicted values. The t-test was used to compare the mean of SCPE to 0 and the Kolmogorov–Smirnov test was used to compare the sampled distribution to the expected one (N(0,1)).

Details on dosing and sampling:
Blood samples (8 mL) were collected from a superficial vein of the forelimbs into heparinized polypropylene tubes immediately before administration and 5, 10, 15, 30, 45, 60 min, and 2, 4, 8, 12, 16, 24 and 36 h after the administration (experiment 1). After collection, blood samples were immediately centrifuged at 2000 x g for 20 min. Plasma was removed and transferred into two polypropylene tubes. Plasma samples were frozen and stored at -30 degree C with quality-control samples prepared in drug-free dog plasma until assay.
Statistics:
Model acceptance: Predicted concentrations (C ) for each individual were plotted versus observed concentrations (C ). Residuals (C –C ) were plotted versus time and versus predicted concentrations.Then, a standardized concentration prediction error (SCPE) was calculated as follows: SCPE = [C obs –C pred ]/SD(C pred ), where SD(C pred) represents the estimated standard deviation on the predicted values. The t-test was used to compare the mean of SCPE to 0 and the Kolmogorov–Smirnov test was used to compare the sampled distribution to the expected one (N(0,1))
Details on distribution in tissues:
Not measured.
Test no.:
#1
Toxicokinetic parameters:
other: Distribution half life (mean): 21.3 +/- 2.1 min
Test no.:
#1
Toxicokinetic parameters:
other: Elmination half life (mean): 3.66 +/- 0.56 hours
Test no.:
#1
Toxicokinetic parameters:
other: Total plasma clearance (mean): 0.72 mL/min/kg
Test no.:
#1
Toxicokinetic parameters:
other: Total clearance (CL) (mean): 0.0430 =/- 0.003 L/h/kg-bw
Test no.:
#1
Toxicokinetic parameters:
other: Two transfer rate constants (k12 and k21) were 0.366 +/- 0.111 (1/h) and 1.55 +/- 0.117 (1/h), respectively.
Test no.:
#1
Toxicokinetic parameters:
other: The initial volume of distribution (Vd) was 0.179 +/- 0.022 L/kg.
Metabolites identified:
not measured
Endpoint:
basic toxicokinetics
Type of information:
experimental study
Adequacy of study:
supporting study
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
comparable to guideline study with acceptable restrictions
Remarks:
Well documented study, similar to OECD 417.
Reason / purpose for cross-reference:
reference to same study
Objective of study:
absorption
Qualifier:
equivalent or similar to guideline
Guideline:
OECD Guideline 417 (Toxicokinetics)
Deviations:
yes
Remarks:
same animals were used to evaluate test substance given orally and via iv
Principles of method if other than guideline:
This study was conducted to determine the pharmacokinetic profile of sodium tungstate after oral administration. Individual pharmacokinetic parameters were estimated using an empirical Bayes' methodology. The linearity of the kinetics was also investigated for different doses administered orally.

Methods similar to methods outlined in "An Application of Population Kinetics Analysis to Estimate Pharmacokinetic Parameters of Sodium Tungstate after Multiple-Dose during Preclinical Studies in Rats." Le Lamer, S. et al. Pharmacology & Technology 2002, 90, 100-105, which indicates methods followed follows "Principles of Laboratory Animal Care" (NIH publication #85-23, revised 1989).
GLP compliance:
not specified
Radiolabelling:
no
Species:
dog
Strain:
Beagle
Sex:
male
Details on test animals or test system and environmental conditions:
TEST ANIMALS
- Source: (Harlan, Grannat, France)
- Weight at study initiation: weighing 11.5 ± 0.24 kg
- Fasting period before study: The dogs were fasted for 24 h before each experiment and were then weighed
- Housing: housed individually during the study in metabolism stainless steel cages
- Individual metabolism cages: yes
- Diet (e.g. ad libitum): access to pelleted food (400 g/day/animal)
- Water (e.g. ad libitum): Tap water was distributed ad libitum
- Acclimation period: The animals underwent an acclimatization period of 10 days before the experiment.

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 18-21 degrees C
- Humidity (%): 40 to 70%,
- Photoperiod (hrs dark / hrs light): artificial lighting, alternating on a 12 h light/dark cycle.



Route of administration:
other: oral gavage and i.v.
Vehicle:
other: distilled water for oral route and 0.9% sterile saline for i.v. route
Details on exposure:
For oral administration, a gavage through a stomach tube using a polypropylene catheter was used.


For iv administration, the dose was administered over 1 minute into the cephalic vein.
Duration and frequency of treatment / exposure:
single oral gavage or i.v. administration
Remarks:
Doses / Concentrations:
25 and 50 mg/kg
No. of animals per sex per dose / concentration:
six animals total
Control animals:
yes
Details on study design:
Tungsten plasma concentrations were determined using an inductively coupled plasma emission spectrometric method.

Individual pharmacokinetic parameters were estimated using an empirical Bayes' methodology. The basic pharmacokinetic parameters considered in the population analysis are clearance (CL), initial volume of distribution (V), transfer rate constants (k12, and k21), absorption rate constant (ka), lag time (tlag), elimination half-time (t1/2elim), and bioavailability (F).




Details on dosing and sampling:
Blood samples (8mL) were collected from a superficial vein of the forelimbs at the following time points 10, 20, 40, and 60 minutes and 2, 4, 8, 12, 16, 24, and 36 hours for oral route. Blood samples (8mL) were collected from a superficial vein of the forelimbs at the following time points 5, 10, 15, 30 , 45, and 60 minutes and 2, 4, 8, 12, 16, 24, and 36 hours for iv route. Blood was also collected before administration of the test substance to establish a baseline.
Statistics:
Results were presented as mean +/- S.D. In dog, a two-way ANOVA was performed to assess the effect of the administered dose on parameters determined by noncompartmental approach (i.e., C max and AUC after oral administration of 25 and 50 mg/kg sodium tungstate). Before the statistical analyses, CL, Vd, Cmax, and AUC were log-transformed; Cmax and AUC were normalized to the same administrated dose. A 5% level of statistical significance were used.
Details on absorption:
The results found in this study indicated that absorption of sodium tungstate when administered in solution form was rapid (tmax was 1-2 hours). The bioavailability averaged 66%. Over the sampling times monitored, plasma concentration profiles versus time were compatible with a two-compartment model and first-order kinetics. The half life in the dog was calculated to be 4 hours.
Toxicokinetic parameters:
Tmax: 1-2 hours
Toxicokinetic parameters:
AUC: 177.0 +/- 53.3 after oral administration of 25 mg/kg dose
Toxicokinetic parameters:
AUC: 431.0 +/- 93.7 after oral administration of 50 mg/kg dose
Toxicokinetic parameters:
other: mean bioavailability was 66% (57-74%)
Toxicokinetic parameters:
half-life 1st: 4 hours
Toxicokinetic parameters:
AUC: 297.2 +/- 34.9 following i.v. administration of 25 mg/kg of test substance
Toxicokinetic parameters:
AUC: 647.4 +/- 70.7 following i.v. administration of 50 mg/kg of test substance
Conclusions:
The results found in this study indicated that absorption of sodium tungstate when administered in solution form was rapid (tmax was 1-2 hours). The bioavailability averaged 66%. Over the sampling times monitored, plasma concentration profiles versus time were compatible with a two-compartment model and first-order kinetics. The half life in the dog was calculated to be 4 hours.
Endpoint:
basic toxicokinetics
Type of information:
experimental study
Adequacy of study:
supporting study
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
comparable to guideline study with acceptable restrictions
Remarks:
Methods well documented study. Similar to OECD 417
Reason / purpose for cross-reference:
reference to same study
Objective of study:
absorption
Qualifier:
equivalent or similar to guideline
Guideline:
OECD Guideline 417 (Toxicokinetics)
GLP compliance:
not specified
Radiolabelling:
no
Species:
rat
Strain:
Sprague-Dawley
Sex:
male
Details on test animals or test system and environmental conditions:
TEST ANIMALS
- Source: IFFA CREDO, L'Arbresle Cedex, France
- Age at study initiation: age 10 weeks
- Weight at study initiation: weighing from 316 to 532 g
- Fasting period before study: Rats fasted overnight (12 h) before drug administration and were then weighed.
- Housing: They were housed in groups in stainless steel cages with suspended wire-mesh floors (maximum of three rats per cage).
- Diet (e.g. ad libitum): The rats were fed a standard laboratory rodent diet (UAR sterile food, Usine d'Alimentation Rationnelle, Villemoisson, Epinay s/Orge, France)
- Water (e.g. ad libitum): allowed free access to drinking water.
- Acclimation period: Animals underwent an acclimatization of a minimum of 2 weeks before treatment

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 18-21 degree C
- Humidity (%): 40 to 70%
- Photoperiod (hrs dark / hrs light): artificial lighting, alternating on a 12 h light/dark cycle.
Route of administration:
other: oral gavage and i.v.
Vehicle:
other: distilled water for oral gavage and 0.9% saline for i.v.
Details on exposure:
Single oral (35.9 and 107.7 mg/kg) doses of sodium tungstate were administered to each rat. Two different solutions of sodium tungstate were prepared on the day of administration, two solutions in distilled water (3.6 and 10.8 mg/mL) for oral administration. These solutions were used to treat animals, under the administered volume of 10 mL/kg. A gavage through a stomach tube using a polypropylene catheter was used.

For iv administration, the dose (8.97 mg/kg) was administered over 1 minute into the cephalic vein.
Duration and frequency of treatment / exposure:
single oral gavage or i.v.
Remarks:
Doses / Concentrations:
35.9 and 107.7 mg/kg orally; and 8.97 mg/kg via i.v.
No. of animals per sex per dose / concentration:
Two hundred sixteen total rats were used. The specific number of rats evaluated per dose per route was not indicated, but one rat per blood sampling event was sacrificed.
Control animals:
yes, concurrent no treatment
Positive control reference chemical:
no
Details on study design:


Tungsten plasma concentrations were determined using an inductively coupled plasma emission spectrometric method

Individual pharmacokinetic parameters were estimated using an empirical Bayes' methodology.

Base structural and statistical models were chosen according to the results of the preliminary analysis performed on the average concentration values at each time point. Thus, the basic pharmacokinetic parameters considered in the population analysis are clearance (CL), initial volume of distribution (V), transfer rate constants (k12, and k21), absorption rate constant (ka), lag time (tlag), elimination half-time (t1/2elim), and bioavailability (F).




Details on dosing and sampling:
In rat, blood samples were collected (one sample per rat) at the following time points (six animals per time point): 5, 10, 15, and 30 min and 1, 2, 4, 8, 12, 16, and 24 h after drug administration. Untreated animals were used for basal tungsten level determination. Two minutes before sampling, rats were anesthetized with diethylether and then sacrificed by section of the carotid artery. Total blood was collected in heparinized polypropylene tubes (0.1 ml sodium heparinate per tube).
Statistics:
Differences in pharmacokinetic parameters were compared across the three treatment groups by using the Kruskal-Wallis test. The effect of the administered dose (35.9 versus 107.7 mg/kg sodium tungstate) was also assessed by comparing AUC.
Details on absorption:
Data were consistent with a two-compartment model. Twelve hours after i.v. administration and 24 h after oral administration of 35.9 mg/kg, concentrations returned to baseline value (i.e., 137 ng/ml). After oral administration, absorption was rapid (approximately 2 h). Bioavailability averaged 92%. Bioavailability was 95% (CV = 9.5%) after oral adminis-tration of 35.9 mg/kg and 88% (CV = 14.8%) after oral administration of 107.7 mg/kg. No dose dependence was observed, suggesting linear kinetics.
Toxicokinetic parameters:
AUC: 26.5 +/- 2.3 following i.v. administration of 8.97 mg/kg
Toxicokinetic parameters:
AUC: 111.2 +/- 10.1 following oral administration of 35.9 mg/kg
Toxicokinetic parameters:
AUC: 326.9 +/- 31.0 following oral administration of 107.7 mg/kg
Toxicokinetic parameters:
other: bioavailability was 95% after oral administration of 35.9 mg/kg
Toxicokinetic parameters:
other: bioavailability was 88% after oral administration of 107.7 mg/kg
Toxicokinetic parameters:
half-life 1st: 1.72 hours following i.v administration of 8.97 mg/kg
Toxicokinetic parameters:
half-life 1st: 1.68 hours following oral administration of 35.9 mg/kg
Toxicokinetic parameters:
half-life 1st: 1.79 hours following oral administration of 107.7 mg/kg
Toxicokinetic parameters:
Tmax: 1-2 hours
Conclusions:
Data were consistent with a two-compartment model. Twelve hours after i.v. administration and 24 h after oral administration of 35.9 mg/kg, concentrations returned to baseline value (i.e., 137 ng/ml). After oral administration, absorption was rapid (approximately 2 h). Bioavailability averaged 92%. Bioavailability was 95% (CV = 9.5%) after oral administration of 35.9 mg/kg and 88% (CV = 14.8%) after oral administration of 107.7 mg/kg. No dose dependence was observed, suggesting linear kinetics.
Endpoint:
basic toxicokinetics
Type of information:
experimental study
Adequacy of study:
supporting study
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
comparable to guideline study with acceptable restrictions
Remarks:
Well documented study, similar to OECD 417
Objective of study:
absorption
Qualifier:
equivalent or similar to guideline
Guideline:
OECD Guideline 417 (Toxicokinetics)
Principles of method if other than guideline:
The purpose of this study was to use a population approach to compute individual pharmacokinetic parameters after repeated oral administrations and to study the influence of the administered dose, gender, and the duration of treatment on the pharmacokinetic parameters.
GLP compliance:
not specified
Radiolabelling:
no
Species:
rat
Strain:
Sprague-Dawley
Sex:
male/female
Details on test animals or test system and environmental conditions:
TEST ANIMALS
- Source: CRIFFA, Barcelona, Spain
- Age at study initiation: aged 10 weeks
- Weight at study initiation: mean weights of males and females were 0.197 and 0.161 kg, respectively
- Housing: The animals were housed in stainless steel Makrolon cages. Each cage contained a maximum of 5 animals of the same sex.
- Diet (e.g. ad libitum): The rats had free access to a pelleted rat diet (UAR sterile food, Usine d'Alimentation Rationnelle, Villemoisson, Epinay s/Orge, France.
- Water (e.g. ad libitum):The water was offered to the animals in bottles ad libitum
- Acclimation period: All animals underwent a period of two weeks of observation and acclimatization before treatment.

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 21-25 degree C
- Humidity (%): 40-65%
- Photoperiod (hrs dark / hrs light): alternating a 12 hr light/dark cycle.
Route of administration:
other: oral gavage and i.v.
Vehicle:
other: distilled water for oral gavage administration and 0.9% sodium chloride for i.v. administration
Details on exposure:
The drug was given in distilled water by a gavage through stomach tubing using a polypropylene catheter.

For intravenous administrations, the dose was administered in an aqueous solution containing 0.9% sodium chloride over 1 min. into the tail vein.
Duration and frequency of treatment / exposure:
Experiment 1: single intravenous administration (dosing was over 1 minute)
Experiment 2: daily oral gavage exposures for 7 days
Experiment 3: daily oral gavage exposures for 8 days
Experiment 4: daily oral gavage exposures for 28 days
Remarks:
Doses / Concentrations:
Experiment 1 (i.v.): 9 mg/kg
Experiment 2 (oral gavage for 7 days): 50, 100, and 200 mg/kg per day
Experiment 3 (oral gavage for 8 days): 50, 100, and 200 mg/kg per day
Experiment 4 (oral gavage for 28 days): 50, 100, and 200 mg/kg per day
No. of animals per sex per dose / concentration:
Experiment 1: 46 male rats
Experiment 2: 54 rats (27 males and 27 females)
Experiment 3: 53 rats (26 males and 27 females)
Experiment 4: 53 rats (26 males and 27 females)
Control animals:
other: blood was taken prior to administration of test substance to obtain a baseline
Details on dosing and sampling:
After intravenous administration (experiment 1), blood samples (one sample per rat) were drawn (6 animals per time-point) before administration (to evaluate basal tungsten levels), then 5, 10, 15, 30 min., and 1, 2, 4, 8, 12, 16 and 24 hr after administration. After multiple oral administrations, blood samples (two samples per rat) were obtained: i) on days 7 and 8 of treatment (experiments 2 and 3, respectively) and ii) on day 28 of treatment (experiment 4), at the following times (6 animals per time-point, 3 males and 3 females): immediately before drug administration and 0.5, 1, 3, 8, and 24 hr after administration. Blood samples were collected in heparinized polypropylene tubes then immediately centrifuged at 3000 x g for 10 min. Plasma was removed and transferred to other tubes, frozen and stored at - 30 degrees, with quality control samples prepared in drug-free rat plasma until assayed.


Tungsten was quantified using an inductively coupled plasma emission spectrometric method.

The basic pharmacokinetic parameters (considered in the population analysis are clearance (CL), initial volume of distribution
(V), transfer rate constants (k12 and k21), absorption rate constant (ka), and bioavailability (F).

Several secondary pharmacokinetic parameters were calculated from the individual primary pharmacokinetic parameters: time of peak
(Tmax), maximum concentration (Cmax), minimum concentration (Cmin), elimination half-time (t1/2 elimination), absorption half-time (t1/2 ka), area under the plasma concentration-time curve (AUC), and volume of distribution (Vd-beta).






Statistics:
Model acceptance: The adequacy of the model to the data was judged by using graphics and descriptive statistics. Individual predicted concentrations (IPRED) were plotted versus observed concentrations (DV) and results were compared to the reference line of slope= 1 and intercept= 0. Moreover, residuals (DV-IPRED) were plotted versus time and versus predicted concentrations. A Standardized Concentration Prediction Error (SCPE) was calculated as follows: SCPE= [DV-IPRED}ISD(IPRED), where SD(IPRED) represents the estimated standard deviation on the predicted values computed using all sources of random variability including the residual error. Then, the Student's t-test was used to compare the mean of SCPE to 0; the Kol-mogorov-Smirnov test was used to compare the sampled distribution to the expected one (N(0,1))
The model was accepted when i) plots showed no systematic pattern and, ii) descriptive statistics did not show any systematic deviation from the initial hypothesis.
Details on absorption:
After repeated oral administrations, the bioavailability F was greater in female rats than in male whatever the administered dose. Our results indicate that the absorption of tungsten was rapid (tmax 1-3 hr) when administered in solution form. Total plasma clearance and elimination half-life averaged 2.8 ml/min./kg and 3.04 hr in males, and 3 ml/min./kg and 2.74 hr in females. The bioavailability averaged 70%; it was significantly higher in females than in males (0.78 versus 0.61). In spite of the fact that the relationship between Cl and the duration of treatment was not retained as a significant explanatory factor in the final model, on the basis of the log-likelihood ratio test, the mean Cmax and AUC were about 30% greater after 28 days of treatment than after 7 and 8 days.
Toxicokinetic parameters:
Tmax: ranged from 2.09 to 2.01 hours (males and females) for the 50 mg/kg dose
Toxicokinetic parameters:
Tmax: ranged from 1.82 to 2.13 hours (males and females) for the 200 mg/kg dose
Toxicokinetic parameters:
Tmax: ranged from 1.87 to 1.67 hours (males and females) for the 200 mg/kg dose
Toxicokinetic parameters:
Cmax: ranged from 10.2 to 12.5 ug//ml (males and females) for the 50 mg/kg dose
Toxicokinetic parameters:
Cmax: ranged from 17.5 to 25.3 ug/ml (males and females) for the 100 mg/kg dose
Toxicokinetic parameters:
Cmax: ranged from 35.8 to 45.6 ug/ml (males and females) for the 200 mg/kg dose
Toxicokinetic parameters:
AUC: ranged from 118.9 to 129.5 mgxhr/l (males and females) for the 50 mg/kg dose
Toxicokinetic parameters:
AUC: ranged from 221.5 to 287.7 mgxhr/l (males and females) for the 100 mg/kg dose
Toxicokinetic parameters:
AUC: ranged from 532.9 to 604.3 mgxhr/l (males and females) for the 200 mg/kg dose
Toxicokinetic parameters:
half-life 1st: ranged from 2.8 to 2.42 hours (males and females) for the 50 mg/kg dose
Toxicokinetic parameters:
half-life 1st: ranged from 2.80 to 2.70 hours (males and females) for the 100 mg/kg dose
Toxicokinetic parameters:
half-life 1st: ranged from 3.47 to 3.10 hours (males and females) for the 200 mg/kg dose

After intravenous administration, a rapid distributive phase was observed (half-life of 14 min.); the two transfer rate constants (k12 and k21) and the initial volume of distribution (V) were 1.38+/-0.046 hr-1, 1.17+/-0.048 hr' and 0.23+/-0.026 1/kg, respectively. CL averaged 0.17+/-0.024 1/hr/ kg and Vd 0.60+/-0.083 I/kg.

Conclusions:
After repeated oral administrations, the bioavailability F was greater in female rats than in male whatever the administered dose. Our results indicate that the absorption of tungsten was rapid (tmax 1-3 hr) when administered in solution form. Total plasma clearance and elimination half-life averaged 2.8 ml/min./kg and 3.04 hr in males, and 3 ml/min./kg and 2.74 hr in females. The bioavailability averaged 70%; it was significantly higher in females than in males (0.78 versus 0.61). In spite of the fact that the relationship between Cl and the duration of treatment was not retained as a significant explanatory factor in the final model, on the basis of the log-likelihood ratio test, the mean Cmax and AUC were about 30% greater after 28 days of treatment than after 7 and 8 days.
Endpoint:
basic toxicokinetics in vivo
Type of information:
experimental study
Adequacy of study:
supporting study
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
comparable to guideline study with acceptable restrictions
Remarks:
Well documented study with sufficient information provided on methods and results to evaluate data.
Objective of study:
absorption
distribution
excretion
Qualifier:
no guideline available
Principles of method if other than guideline:
An empirical Bayes' methodology was used to determine the pharmacokinetic profile of sodium tungstate in beagle dogs after multiple oral dosing using the P-PHARM computer program.
GLP compliance:
not specified
Radiolabelling:
no
Species:
dog
Strain:
Beagle
Sex:
male/female
Details on test animals or test system and environmental conditions:
TEST ANIMALS
- Source: Harlan (Grannat, France)
- Weight at study initiation: Mean weight 10.5 kg
- Housing: housed individually during the study in stainless steel cages
- Diet (e.g. ad libitum): access to pelleted food (400 g/day per animal); Usine d’Alimentation Rationnelle, Villemoisson sur Orge, France)
- Water (e.g. ad libitum): Tap water was distributed ad libitum.
- Acclimation period: The animals underwent an acclimatization period of 10 days before the experiment.

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 18–21degree C
- Humidity (%): 40–70%
- Photoperiod (hrs dark / hrs light): constant 12 h light/dark cycle (lights on at 08.00 h)- artificial lighting
Route of administration:
oral: gavage
Vehicle:
other: Distilled water
Details on exposure:
For oral administration, Sodium tungstate was administered to animals in a solution of distilled water. The drug was given by a gavage through stomach tubing using a polypropylene catheter.

Duration and frequency of treatment / exposure:
- 18 dogs (nine males and nine females) received repeated oral doses of (7 and 14 mg/kg three times a day) for 11 days (experiment 2)
- 28 dogs (14 males and 14 females) received repeated oral doses of sodium tungstate (5, 10 and 20 mg/kg three times a day) for 80 days (experiment 3).
Remarks:
Doses / Concentrations:
5, 7, 10, 14 and 20 mg/kg
No. of animals per sex per dose / concentration:
- 18 dogs (nine males and nine females) received repeated oral doses of (7 and 14 mg/kg three times a day)
- 28 dogs (14 males and 14 females) received repeated oral doses of sodium tungstate (5, 10 and 20 mg/kg three times a day)
Control animals:
no
Positive control reference chemical:
no
Details on study design:
During the treatment period, each dog was observed daily on at least two occasions and the fluid and food intake were measured. The animals were weighed twice a week in order to calculate daily doses. Laboratory studies including hematology, blood chemistry, and urinalysis were performed at baseline, after 6 weeks of treatment, and at the end of treatment.

Tungsten was quantified using an inductively coupled plasma emission spectrometric method at a wavelength of 207.91 nm. Calibration curves were obtained in the range 134–1300 ng/mL. Precision ranged from 0.4 to 17%, and accuracy was between 89 and 105%. Dilution has no influence on the performance of the method, which could then be used to quantify plasma containing up to 90 mg/mL. The limit of quantification was 100 ng/mL (precision, 17%). Plasma concentrations less than the limit of quantitation were omitted from the analysis.

Individual pharmacokinetic parameters were estimated using an empirical Bayes methodology. The population analysis was performed using the P-PHARM computer program (version 1.4, SIMED, Cre´teil, France). The population estimation algorithm is an EM-type procedure. The kinetic parameters considered in the population analysis consists of total clearance (CL), initial volume of distribution (V), transfer rate constants (k12 and k21 ), absorption rate constant (ka ), lag-time (tlag ) and bioavailability (F). Several secondary pharmacokinetic parameters were calculated from the individual (Bayesian estimates) primary pharmacokinetic parameters: the model-predicted maximum concentration (C ) and the time of peak (tmax), the area under the plasma concentration–time curve (AUC); the elimination half-life (t1/2elim ); and the of distribution (Vd-β). Moreover, the plasma concentration of tungsten at any time can be easily predicted using the empirical Bayes estimate of the pharmacokinetic parameters.

To determine model acceptance, predicted concentrations (Cpred ) for each individual were plotted versus observed concentrations (Cobs ). Residuals (Cpred –C obs) were plotted versus time and versus predicted concentrations. Then, a standardized concentration prediction error (SCPE) was calculated as follows: SCPE = [Cobs -Cpred]/SD(Cpred), where SD(Cpred ) represents the estimated standard deviation on the predicted values. The t-test was used to compare the mean of SCPE to 0 and the Kolmogorov–Smirnov test was used to compare the sampled distribution to the expected one (N(0,1)).

Details on dosing and sampling:
Blood samples (8 mL) were collected from a superficial vein of the forelimbs into heparinized polypropylene tubes on day 11 of treatment, immediately before drug administration and 0.25, 0.5, 1, 2, 4, and 8 h postdose (experiment 2); on days 8 and 80 of treatment, immediately before drug administration and 0.25, 0.5, 1, 2, 4, and 8 h postdose (experiment 3). After collection, blood samples were immediately centrifuged at 2000 x g for 20 min. Plasma was removed and transferred into two polypropylene tubes. Plasma samples were frozen and stored at -30 degree C with quality-control samples prepared in drug-free dog plasma until assay.
Statistics:
Model acceptance: Predicted concentrations (C ) for each individual were plotted versus observed concentrations (C ). Residuals (C –C ) were plotted versus time and versus predicted concentrations.Then, a standardized concentration prediction error (SCPE) was calculated as follows: SCPE = [C obs –C pred ]/SD(C pred ), where SD(C pred) represents the estimated standard deviation on the predicted values. The t-test was used to compare the mean of SCPE to 0 and the Kolmogorov–Smirnov test was used to compare the sampled distribution to the expected one (N(0,1))
Test no.:
#2
Toxicokinetic parameters:
Tmax: (mean) 1.91-2.14 hours for doses ranging from 21-42 mg/kg bw/day
Test no.:
#3
Toxicokinetic parameters:
Tmax: (mean) 1.78-2.05 hours for doses ranging from 15-60 mg/kg bw/day
Test no.:
#2
Toxicokinetic parameters:
AUC: (mean) 46.6 and 78.3 mg/L*h for the 21 and 42 mg/kg bw/day dose groups
Test no.:
#3
Toxicokinetic parameters:
AUC: (mean) 35.7, 72.0 and 148.1 mg/L*h for the 15, 30 and 60 mg/kg bw/day dose groups
Test no.:
#2
Toxicokinetic parameters:
Cmax: (mean) 9.5 and 17.3 ug/mL for the 21 and 42 mg/kg bw/day dose groups, respectively.
Test no.:
#3
Toxicokinetic parameters:
Cmax: (mean) 6.98, 14.3 and 27.7 ug/mL for the 15, 30 and 60 mg/kg bw/day dose groups, respectively.
Test no.:
#2
Toxicokinetic parameters:
other: Elimination half life (mean): 2.71-2.84 hours for doses ranging from 21-42 mg/kg bw/day
Test no.:
#3
Toxicokinetic parameters:
other: Elimination half life (mean): 2.60-2.67 hours for doses ranging from 15-60 mg/kg bw/day
Test no.:
#2
Toxicokinetic parameters:
other: Mean bioavailability (F) ranged from 45-53% for doses ranging from 21-42 mg/kg bw/day
Test no.:
#3
Toxicokinetic parameters:
other: Mean bioavailability (F) ranged from 57-59% for doses ranging from 15-60 mg/kg bw/day
Test no.:
#2
Toxicokinetic parameters:
other: Total clearance (CL): 0.0448-0.0450 L/h/kg for doses ranging from 21-42 mg/kg bw/day
Test no.:
#3
Toxicokinetic parameters:
other: Total clearance (CL): 0.0445-0.0448 L/h/kg for doses ranging from 15-60 mg/kg bw/day
Test no.:
#2
Toxicokinetic parameters:
other: Elimination half life (mean): 2.71-2.84 h for doses ranging from 21-42 mg/kg bw/day
Test no.:
#3
Toxicokinetic parameters:
other: Elimination half life (mean): 2.60-2.67 hrs for doses ranging from 15-60 mg/kg bw/day
Metabolites identified:
not measured

Although bioavailability, AUC, and elimination half life were higher in male than female, the introduction of gender into the final model did not contribute to a statistical improvement of the fit of the population pharmacokinetic model. In the range 15 -60 mg/kg/day sodium tungstate, the treatment duration did not change the pharmacokinetic parameters.

At 60 mg/kg per day, a decrease in the mean lymphocyte count of 1.5-fold, compared to baseline, was observed and a 2-fold increase in total cholesterol occurred. Although no vomiting was observed the days of kinetics, some episodic vomiting was observed in about 10% of the animals.

Conclusions:
The results indicate that after repeated oral administration (11 days or 13 weeks) of 5–20 mg/kg sodium tungstate three times a day in dogs, tungsten plasma concentrations increased in proportion to dose. No dose-dependent changes in pharmacokinetic parameters were observed.

Description of key information

A number of high quality toxicokinetic studies were available on sodium tungstate on mice, rats, and dogs administered via the oral and inhalation routes. Following oral administration of sodium tungstate in mice, there was a slight increase in tungsten tissue concentration for treated rodents (vs. endogenous concentrations-control) in the plasma, intestines, liver, and femur after 14 days. No difference in concentration was observed in the kidneys. In contrast, the uterus displayed a large increase in tungsten concentration and was significantly above endogenous (control) levels. Following oral administration of sodium tungstate in rats, absorption was rapid (1-3 hours), with the bioavailability ranging from 70 to 92%. There was an observed increase in tungsten tissue concentration for treated rodents (vs. endogenous concentrations-control) in plasma, and all tissues examined. The plasma, liver, and uterus showed tungsten levels only slightly above baseline levels; however, the intestine, kidneys, and femur displayed significant increases in the amount of tungsten. The half life in rats was calculated to range from around 1-3 hours. Following oral administration of sodium tungstate in dogs, the absorption was rapid (Tmax was 1-2 hours), with the bioavailability averaging 66%. Plasma concentration profiles versus time were compatible with a two-compartment model and first-order kinetics. The half life in the dog was calculated to be 4 hours. Following inhalation administration of sodium tungstate to rats, accumulation varied among different tissues with the highest concentration seen in the lung, nasal epithelium, thyroid, femur, and kidney. Tissues with minimal accumulation included lung lymph nodes, heart, thymus, adrenal gland, testes, pancreas, and spleen. Levels of tungsten observed in all brain structures examined paralleled blood concentrations at 2-3% of blood levels and were highest at the end of exposure, returning to background level within 3 days. The highest concentrations of tungsten in the brain were seen in the pituitary gland where levels of tungsten seen in the olfactory bulb, striatum, cerebellum, and cortex were much lower suggesting tungsten is largely excluded by the blood brain barrier. Urine concentrations of radio-labeled tungsten were elevated throughout the first 7 days post exposure. Tungsten elimination from most tissue compartments was rapid. Tissues with slower terminal elimination rates include femur, lung, and spleen. The available toxicokinetic data on sodium tungstate indicate that it is readily bioavailable via the oral route of administration, and that by this route tungsten accumulates primarily in the intestines, kidneys, and femur. When administered via inhalation, tungsten primarily accumulated in the lung, nasal epithelium, thyroid, femur, and kidney. The half-life of tungsten following administration of sodium tungstate ranged from 1-4 hours, which indicates that it is rapidly excreted.

Key value for chemical safety assessment

Bioaccumulation potential:
no bioaccumulation potential

Additional information