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Basic toxicokinetics

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basic toxicokinetics
Type of information:
experimental study
Adequacy of study:
key study
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
comparable to guideline study with acceptable restrictions
Similar to OECD Test Guideline 417 for toxicokinetics-distribution studies with acceptable deviations.
Reason / purpose for cross-reference:
reference to same study

Data source

Reference Type:
Distribution of Tungstate in Pregnant Mice and Effects on Embryonic Cells in Vitro
Wide, M., Danielsson, B.R.G., and Dencker, L.
Bibliographic source:
Environmental Research 40, 487-498

Materials and methods

Objective of study:
other: retention
Test guideline
equivalent or similar to guideline
OECD Guideline 417 (Toxicokinetics)
Version / remarks:
: Section on Distribution Studies
GLP compliance:

Test material

Constituent 1
Chemical structure
Reference substance name:
Disodium wolframate
EC Number:
EC Name:
Disodium wolframate
Cas Number:
Molecular formula:
disodium dioxotungstenbis(olate)
Details on test material:
- Source: purchased from Amersham International plc
- Physical form: sodium tungstate in a dilute sodium hydroxide solution
[185W] tungstate

Test animals

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

Administration / exposure

Route of administration:
other: injection
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.
Doses / concentrations
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.
- 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).

Results and discussion

Toxicokinetic / pharmacokinetic studies

Details on distribution in tissues:
- 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.

Applicant's summary and conclusion

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.