Vanadium dioxide

Endpoint summary

Description of key information

A comprehensive literature search was recently conducted for the vanadium category substances, to source relevant information for the hazard and risk assessment. For the group of the soluble vanadium substances, a limited number of studies is available and the different experimental approaches lead to a variety of endpoints measured.

Of the limited effects noted following oral exposure with soluble vanadium substances, it appears most likely that effects on haematological parameters are the most consistently reported among a number of investigators (Mountain et al 1953, Zaporowska et al. 1993, Scibior et al 2006, Scibior, 2005, NTP, 2002). Altogether, haematological effects have been found with a variety of different vanadium compounds including sodium metavanadate, vanadium pentoxide, and ammonium metavanadate supporting the use of this endpoint for risk assessment purposes.

Key value for chemical safety assessment

Repeated dose toxicity: via oral route - systemic effects

Link to relevant study records

Reference

Reference 1

Endpoint:

short-term repeated dose toxicity: oral

Type of information:

experimental study

Adequacy of study:

supporting study

Reliability:

3 (not reliable)

Rationale for reliability incl. deficiencies:

significant methodological deficiencies

Remarks:

Well reported study, however, the relevance of the study is limited because only selected parameters were investigated and animals were dosed for only 4 weeks. The test material was insufficiently described, body weight data were recorded but not reported, haematology, clinical biochemistry, FOB, necropsy, ophthalmological examination, thyroid hormones, organ weights and histopathology were not conducted/evaluated, applied doses were not analytically analysed, results were reported only in figures and raw data were not provided, historical control data were not provided either.

Qualifier:

no guideline followed

Principles of method if other than guideline:

Two month old Wistar rats of both sexes received an aqueous solution of ammonium metavanadate (AMV) at a concentration of 0.01, 0.05, 0.15 and 0.30 mg V/mL (corresponding to 0.2, 1, 3 and 6 mM solution) over 4 weeks. Food and water intake was monitored daily, body weight was recorded weekly. At study termination all animals were sacrificed and liver, kidney, adrenals and spleen were isolated for L-ascorbic acid determination.

GLP compliance:

not specified

Limit test:

no

Species:

rat

Strain:

Wistar

Remarks:

albino

Sex:

male/female

Details on test animals or test system and environmental conditions:

- Age at study start: two month
- Diet: standard granulated roden laboratory chow- LSM (CLPP, Motycz, Poland)
- Water: control group received deionized drinking water ad libitum, treatment groups received AMV containing drinking water ad libitum
- Housing: animals were individually housed in stainless steel cages under controlled conventional conditions

ENVIRONMENTAL CONDITIONS
- Temperature: 19-20 °C
- Humidity: 60±10 %
- Photoperiod: natural day-night cycle

Route of administration:

oral: drinking water

Vehicle:

other:

Remarks:

deionized drinking water

Analytical verification of doses or concentrations:

not specified

Duration of treatment / exposure:

4 weeks

Frequency of treatment:

drinking water, ad libitum

Dose / conc.:

0.01 other: mg/mL

Remarks:

males: 1.18±0.01 mg/kg bw/day
females: 1.50±0.23 mg/kg bw/day
(calculated based on ingested amount of AMV solution)

Dose / conc.:

0.05 other: mg/mL

Remarks:

males: 4.93±0.15 mg/kg bw/day
females: 6.65±0.27 mg/kg bw/day
(calculated based on ingested amount of AMV solution)

Dose / conc.:

0.15 other: mg/mL

Remarks:

males: 12.99±1.42 mg/kg bw/day
females: 13.38±1.79 mg/kg bw/day
(calculated based on ingested amount of AMV solution)

Dose / conc.:

0.3 other: mg/mL

Remarks:

males: 22.06±1.29 mg/kg bw/day
females: 26.62±1.41 mg/kg bw/day
(calculated based on ingested amount of AMV solution)

No. of animals per sex per dose:

males:
group 1: 20, group 2: 15, group 3: 16, group 4: 11, group 5: 15
females:
group 1: 20, group 2: 15, group 3: 16, group 4: 13, group 5: 15

Control animals:

yes

Details on study design:

- Rationale for animal assignment: animals were randomly divided into 5 groups

Positive control:

no data

Observations and examinations performed and frequency:

BODY WEIGHT: Yes
- Time schedule for examinations: weekly

FOOD CONSUMPTION:
- Food consumption for each animal determined:Yes
- Time schedule for examinations: daily

WATER CONSUMPTION AND COMPOUND INTAKE: Yes
- Time schedule for examinations: daily
The vanadium intake was calculated on the basis of the amount of AMV solution consumed by the rats.

Other examinations:

Determination of L-Ascorbic acid concentration in liver, adrenals, spleen and kidney
All organs were immediately washed with ice-cold phosphate buffered saline (PBS; pH 7.4) and delicate by desiccated with lignin. Then the portions of internal organs were homogenized in ice-cold 0.25 M sucrose. The homogenates (5%; w/v) were centrifuged at 3000 x g 10 min at 4°C. In the obtained supernatant L-ascorbic acid concentration was measured according to the method of Kyaw (1978). All assays were duplicated.

Statistics:

Student's t-test was used for statistical analysis. A value of P < 0.05 was used as the level significance. All results are presented as mean values_ SEM.

Clinical signs:

not examined

Mortality:

not examined

Body weight and weight changes:

not specified

Food consumption and compound intake (if feeding study):

effects observed, treatment-related

Description (incidence and severity):

In animals receiving as sole drinking liquid an aqueous AMV solution of 0.15 and 0.30 mg V/mL concentration, a statistically significant and dose dependent decrease of food uptake was observed as compared with the control.
For details please refer to the field "attached background material".

Food efficiency:

not examined

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

effects observed, treatment-related

Description (incidence and severity):

In animals receiving as sole drinking liquid an aqueous AMV solution of 0.15 and 0.30 mg V/mL concentration, a statistically significant and dose dependent decrease of AMV solution uptake was observed as compared with the control.
For details please refer to the field "attached background material".

Ophthalmological findings:

not examined

Haematological findings:

not examined

Clinical biochemistry findings:

not examined

Urinalysis findings:

not examined

Behaviour (functional findings):

not examined

Immunological findings:

not examined

Organ weight findings including organ / body weight ratios:

not examined

Gross pathological findings:

not examined

Neuropathological findings:

not examined

Histopathological findings: non-neoplastic:

not examined

Histopathological findings: neoplastic:

not examined

Other effects:

not examined

Details on results:

L-ascorbic acid concentration in tissues:
In animals of both sexes receiving as the only drinking fluid the AMV solutions, a distinct tendency of the L-ascorbic acid level to decrease was noted in the liver, kidneys, spleen and adrenals.
For details please refer to the field "attached background material".

Dose descriptor:

NOEL

Effect level:

0.3 other: mg/mL

Based on:

element

Sex:

male/female

Basis for effect level:

other: no adverse effect observed

Dose descriptor:

NOEL

Effect level:

22.06 mg/kg bw/day (nominal)

Based on:

element

Sex:

male

Basis for effect level:

other: see remarks

Dose descriptor:

NOEL

Effect level:

26.62 mg/kg bw/day (nominal)

Based on:

element

Sex:

female

Basis for effect level:

other: no adverse effect observed

Conclusions:

As reported in several other studies, effects on palatability of vanadium containing fluids were observed in high dose groups. Significant and dose-dependent decreases were observed in L-ascorbic acid levels in spleen, liver, adrenals and kidneys in males exposed to 0.15 and 0.30 mg V/mL and females exposed to 0.05, 0.15 and 0.3 mg V/mL. However, as the adversity of this effect remains questionable (discussed below) and no systemic toxicity was observed, this effect is not considered to be adverse and thus not taken into account for NOAEL setting. Based on this, 0.30 mg V/mL (equivalent to 22.06 mg V/kg bw/day in males and 26.62 mg V/kg bw/day in females) represents the NOEL.

This study is well reported, however, no guideline was followed and only limited parameters were analysed. Significant and dose-dependent decreases were observed in L-ascorbic acid levels in spleen, liver, adrenals and kidneys in males exposed to 0.15 and 0.30 mg V/mL and females exposed to 0.05, 0.15 and 0.3 mg V/mL. Zaporwoska et al. (1993) reported already one year before that L-ascorbic acid levels were decreased in plasma and erythrocytes in rats exposed to ammonium metavanadate. Thus, exposure to ammonium metavanadate seems to be correlated with decreased L-ascorbic acid values in different tissues and body fluids. However, as no systemic toxicity or any other effect was observed, it remains unclear whether mild to moderate L-ascorbic acid depression in tissues is or results in any adverse effects. In a publication of Chan & Reade (1996) Wistar Shionogi rats, unable to synthesize L-ascorbic acid, were supplemented with different doses of L-ascorbic acid to determine the L-ascorbic acid requirements in Wistar rats. After 26 weeks, all animals survived and showed no clinical signs of scurvy. The average weekly body weight gain was normal. A severe L-ascorbic acid deficiency would include perinasal and peri-and intra-oral haemorrhage, joint or intramuscular haemorrhage, weakened or fractured hind limbs, delayed wound healing and a failure to thrive (Clemetson, 1989). As none of these clinical signs were observed in studies reported by Zaporowska (1993, 1994) or Chan & Reade (1996), it is assumed that this mild to moderate L-ascorbic acid depression observed in this study is not an adverse effect.

Apart from that, it has been demonstrated that the toxicity of vanadium increases with its valency. Thus, compounds containing 5 -valent V, such as ammonium metavanadate, are most poisonous. It is known that 5 -valent V enters cells through anion channels, i.e. phosphate or sulfate channels. In cells, 5 -valent V is reduced to 2 -valent VO by some reducing compounds such as L-ascorbic acid and thiol-containing cysteine. Thus, the reported reduction of L-ascorbic acid in several organs and also in blood (Zaporowska et al. 1993) is most likely the result of an enhanced consumption/reduction activity of this compound. However, in contrast to humans, rodents are able to synthesize L-ascorbic acid. Thus, it can be assumed that this reduction of L-ascorbic acid will induce re-synthesis and reduction of L-ascorbic acid is an adaptive but not an adverse effect. Apart from that, it is also noteworthy that humans are not able to synthesize L-ascorbic acid. Thus, it is assumed that the protective reduction of 5 -valent V to 2 -valent VO is performed by another reducing substance and thus, it remains questionable whether a reduction of L-ascorbic acid in rats is relevant for humans.

References:
Clemetson, I.B. et al. (1975): Synthesis and some major functions of vitamin C in aniamsl. Annals of New York Academy of Science 258, 24 -46

Executive summary:

Two-month old Wistar rats of both sexes received an aqueous solution of ammonium metavanadate (AMV) at a concentration of 0.01, 0.05, 0.15 and 0.30 mg V/mL (corresponding to 0.2, 1, 3 and 6 mM solution) over 4 weeks. Food and water intake were monitored daily, body weight was recorded weekly. At study termination all animals were sacrificed, and liver, kidney, adrenals and spleen were isolated for L-ascorbic acid determination.

According to the author the food and fluid consumption was significantly decreased in males and females exposed to 0.15 and 0.30 mg V/mL drinking water. Additionally, the L-ascorbic acid levels in liver, adrenals, spleen and kidneys were significantly and dose-dependently decreased in males and females dosed with 0.05, 0.15 and 0.30 mg V/mL drinking water.

 

Reference 2

Endpoint:

sub-chronic toxicity: oral

Type of information:

experimental study

Adequacy of study:

weight of evidence

Study period:

no data

Reliability:

3 (not reliable)

Rationale for reliability incl. deficiencies:

significant methodological deficiencies

Remarks:

Well document study, however, beside general toxicity, the investigations were restricted to liver and renal function parameters and histopathology. The following deficiencies were noted: test item insufficiently described, no analytical verification of applied doses, only males were used, haematology missing, clinical biochemistry parameter missing, missing organ weights, several organs were not histopathologically examined, details on gross pathology not given, ophthalmological examination not performed, FOB not conducted, thyroid hormones not determined, historical control data missing and individual/raw data not provided, details on histopathological findings were not provided.

Qualifier:

no guideline followed

Principles of method if other than guideline:

Sodium metavanadate was given to groups of male Sprague-Dawley rats at concentrations of 0, 5, 10 and 50 ppm for three months. Liver and renal function parameters were determined in blood at the end of exposure period and organ weights were taken (liver, kidneys, heart, spleen, lung). Heart, liver, lungs, kidneys, spleen, stomach, small and large intestine were histopathologically examined in three rats of each group.

GLP compliance:

not specified

Limit test:

no

Species:

rat

Strain:

Sprague-Dawley

Sex:

male

Details on test animals or test system and environmental conditions:

TEST ANIMALS (40 males)
- Source: Biocentre (Barcelona, Spain)
- Weight at study initiation: about 91.6 +/- 10.8 g at the start of exposure
- Housing: all animals were placed in individual metabolism cages
- Diet: ad libitum (perfectly balanced Panlab diet, Barcelona, Spain), neglible vanadium concentration
- Water: ad libitum

Route of administration:

oral: drinking water

Vehicle:

water

Details on oral exposure:

PREPARATION OF DOSING SOLUTIONS:
- Solutions of sodium metavandate and water were prepared without heating at pH 7.4 and stored at 21-24 °C

Analytical verification of doses or concentrations:

not specified

Details on analytical verification of doses or concentrations:

no data

Duration of treatment / exposure:

3 months

Frequency of treatment:

continuous (in drinking water)

Dose / conc.:

5 ppm

Remarks:

nominal in water

Dose / conc.:

10 ppm

Remarks:

nominal in water

Dose / conc.:

50 ppm

Remarks:

nominal in water

No. of animals per sex per dose:

10 males per group

Control animals:

yes, concurrent vehicle

Details on study design:

no data

Positive control:

no data

Observations and examinations performed and frequency:

CAGE SIDE OBSERVATIONS: Yes
- Time schedule: no data

DETAILED CLINICAL OBSERVATIONS: No data

BODY WEIGHT: Yes
- Time schedule for examinations: weekly

FOOD CONSUMPTION: Yes
- Food consumption for each animal determined and mean daily diet consumption calculated as g food/kg body weight/day: Yes, measured daily

FOOD EFFICIENCY: Yes
- Body weight gain in kg/food consumption in kg per unit time X 100 calculated as time-weighted averages from the consumption and body weight gain data: protein efficiency coefficient was calculated weekly

WATER CONSUMPTION AND COMPOUND INTAKE (if drinking water study): Yes
- Time schedule for examinations: daily

OPHTHALMOSCOPIC EXAMINATION: No data

HAEMATOLOGY: No data

CLINICAL CHEMISTRY: Yes,
- Time schedule for collection of blood: at the end of exposure
- How many animals: 5 animals per group
- Parameters checked: serum analyses was carried out (GOT, GPT, total protein, bilirubin, urea, uric acid,creatine, glucose and cholesterol)

URINALYSIS: Yes
- Metabolism cages used for collection of urine: Yes
- Parameters checked: volume excreted

NEUROBEHAVIOURAL EXAMINATION: No data

IMMUNOLOGY: Not specified

OTHER
- Vanadium concentration in liver, kidneys, spleen, heart and lungs were determined

Sacrifice and pathology:

All animals not used for clinical chemistry were killed at the end of exposure.

GROSS PATHOLOGY: Yes
- Necropsy was performed on all animals after exsanguination.
- Organ weights of liver, kidneys, heart, spleen and lungs were determined.

HISTOPATHOLOGY: Yes:
- Histological examination of heart, lungs, liver, kidneys, spleen, stomach and small and large intestine in three rats of each group.

Other examinations:

no

Statistics:

The significance of differences between control and treated groups was calculated by the Student's test. A difference is considered to be significant when p<0.05.

Clinical signs:

no effects observed

Mortality:

no mortality observed

Body weight and weight changes:

no effects observed

Food consumption and compound intake (if feeding study):

no effects observed

Food efficiency:

no effects observed

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

no effects observed

Ophthalmological findings:

not specified

Haematological findings:

not specified

Clinical biochemistry findings:

effects observed, treatment-related

Description (incidence and severity):

- The amount of total protein was significantly increased in the highest dose group.
- The plasma concentrations of urea and uric acid were increased in the highest exposure group.

Urinalysis findings:

no effects observed

Description (incidence and severity):

on volume of urine

Behaviour (functional findings):

not specified

Immunological findings:

not specified

Organ weight findings including organ / body weight ratios:

no effects observed

Gross pathological findings:

not specified

Neuropathological findings:

not specified

Histopathological findings: non-neoplastic:

effects observed, treatment-related

Description (incidence and severity):

- Histopathological investigation showed mild, though dose-dependent, lesions in kidneys, lungs and spleen in all dosage groups.
- The changes were more evident in the highest dose group (no further details presented).
- Hyperthrophy and hyperplasia were observed in the white pulp of spleen.
- Corticomedullar micro haemorrhagic foci were seen in kidneys.
- Mononuclear cell infiltration, mostly perivascular, were found in lungs.

Histopathological findings: neoplastic:

not specified

Other effects:

not specified

Details on results:

CLINICAL SIGNS AND MORTALITY
- Appearance, behaviour and mortality of the treated rats of all groups were not affected.

BODY WEIGHT AND WEIGHT GAIN
- Growth of the treated rats of all groups was not affected.

FOOD CONSUMPTION AND COMPOUND INTAKE (if feeding study)
- Food consumption of the treated rats of all groups was not affected.

WATER CONSUMPTION AND COMPOUND INTAKE (if drinking water study)
- Water consumption of the treated rats of all groups was not affected.

CLINICAL CHEMISTRY
- Enzyme activities (GOT, GPT) and the amounts of bilirubin in the plasma did not indicate effects on the liver function.
- Glucose and cholesterol showed no changes.

URINALYSIS
- No effects on volume of urine.

ORGAN WEIGHTS
- No dose-dependent differences between treated animals and controls.

OTHER FINDINGS
- Vanadium accumulation was first observed at the dose of 10 ppm in the kidneys and spleen and increased dose-dependently.
- At the highest dose, enrichment of V was also observed in liver, heart and lungs compared to the lower doses and the controls.

Dose descriptor:

NOEL

Effect level:

1.51 mg/kg bw/day (nominal)

Based on:

test mat.

Sex:

male

Basis for effect level:

histopathology: non-neoplastic

Dose descriptor:

NOEL

Effect level:

0.62 mg/kg bw/day (nominal)

Based on:

element

Sex:

male

Basis for effect level:

histopathology: non-neoplastic

Critical effects observed:

not specified

Conversion from ppm (mg/L) to mg/kg bw /d based on:

Study Duration: three months/91 d

Average drinking water consumption): 3747.3  mL/rat   => 41.2 ml/rat/d

Average body weight: [91.6 (start) + 450.2 (end)]/2 = 272 g (divided by two to use as mean weight during the study)

10 mg NaVO3/L = 1.51 mg/kg bw/d

50 mg NaVO3/L = 7.57 mg/kg bw/d

Conclusions:

Sodium metavanadate in drinking water was given to four groups, each consisting of 10 male Sprague-Dawley rats, at concentrations of 0, 5, 10 and 50 ppm. Liver and renal function parameters were determined in blood at the end of exposure period and organ weights were taken (liver, kidneys, heart, spleen, lung). Heart, liver, lungs, kidneys, spleen, stomach, small and large intestine were histopathologically examined in three rats of each group.
Oral administration of NaVO3 via drinking water to groups of male rats over 3 months at concentrations of 0, 5, 10 and 50 ppm caused mild, dose-dependent lesions in kidneys, lungs and spleen with the highest incidence in the 50 ppm group, and increased plasma concentrations of protein, urea and uric acid in the high dose group. Thus, the highest dose level (7.57 mg/kg bw/d NaVO3) represents a clear LOAEL, and the mid dose level (1.51 mg/kg bw/d NaVO3) represents a NOAEL.

Reference 3

Endpoint:

chronic toxicity: oral

Type of information:

experimental study

Adequacy of study:

weight of evidence

Study period:

no data

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

comparable to guideline study with acceptable restrictions

Remarks:

Well documented study. However, the following deficiencies were noted: test substance insufficiently described, low group sizes (n=8), missing parameters in clinical biochemistry and haematology, FOB was not conducted, only one sex was used, individual body weight data, ophthalmological examination missing, thyroid hormones were not determined, applied doses were not analytically verified, some organ weights were missing and not the complete table of organs were histopathologically examined. Historical control data were not provided and most of the data were presented as graphical overview, thus no raw/individual data were provided. The dose setting was changed during the study.

Qualifier:

no guideline followed

Principles of method if other than guideline:

This short description is restricted to non-diabetic rats. Investigations on the diabetic rats are not considered here. Four groups of non-diabetic male rats received different concentrations of VOSO4 in drinking water for 52 weeks. The low dose group received 500 mg/L for 52 weeks. The mid dose group received 500 mg/L for 1 week and then 750 mg/L for 51 weeks. The high dose group received 500 and 750 mg/L for 1 week each and then 1250 mg/L for further 50 weeks. Subgroups of 3 animals from each of the four groups were followed for further 16 weeks after cessation of VOSO4 treatment. Various parameters were examined and determined.

GLP compliance:

not specified

Limit test:

no

Species:

rat

Strain:

Wistar

Sex:

male

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Charles River, Montreal, Quebec, Canada
- Weight at study initiation: 270-320 g
- Diet (ad libitum): standard laboratory food (Purina rat chow)
- Water: ad libitum: vanadyl sulphate hydrate mixed with tap water

ENVIRONMENTAL CONDITIONS
- Photoperiod (hrs dark / hrs light): 12/12

Route of administration:

oral: drinking water

Vehicle:

water

Details on oral exposure:

PREPARATION OF DOSING SOLUTIONS:
- Vanadyl sulphate hydrate (Fischer Scientific Co., Lair lawn, NJ, USA) was mixed with drinking water.
- Solutions were prepared on alternative days.

Analytical verification of doses or concentrations:

not specified

Details on analytical verification of doses or concentrations:

no data

Duration of treatment / exposure:

52 weeks

Frequency of treatment:

continuously (ad libitum in drinking water)

Dose / conc.:

500 mg/L drinking water

Remarks:

for 52 weeks (low dose group); nominal in water

Dose / conc.:

500 mg/L drinking water

Remarks:

500 mg/L for 1 week (mid dose group); nominal in water

Dose / conc.:

750 mg/L drinking water

Remarks:

for 51 weeks (mid dose group); nominal in water

Dose / conc.:

500 mg/L drinking water

Remarks:

for 1 week (high dose group); nominal in water

Dose / conc.:

750 mg/L drinking water

Remarks:

for 1 week (high dose group); nominal in water

Dose / conc.:

1 250 mg/L drinking water

Remarks:

for 50 weeks (high dose group); nominal in water

No. of animals per sex per dose:

8 male rats per group

Control animals:

yes

Details on study design:

- Rationale for animal assignment: random

Positive control:

no data

Observations and examinations performed and frequency:

CAGE SIDE OBSERVATIONS: Yes
- Time schedule: animals were examined closely for possible occurrance of diarrhoea, cataract formation or deterioration of their general condition.

DETAILED CLINICAL OBSERVATIONS: No data

BODY WEIGHT: Yes
- Time schedule for examinations: every 3-5 weeks; every 2-3 weeks during post-exposure observation period.

FOOD CONSUMPTION: Yes
- Food consumption for each animal determined and mean daily diet consumption calculated as g food/kg body weight/day: measured every 3-5 weeks; every 2-3 weeks during post-exposure observation period.

FOOD EFFICIENCY: No data

WATER CONSUMPTION AND COMPOUND INTAKE (if drinking water study): Yes
- Time schedule for examinations: regularly, every 3-5 weeks; every 2-3 weeks during post-exposure observation period

OPHTHALMOSCOPIC EXAMINATION: No

HAEMATOLOGY: Yes
- Time schedule for collection of blood: hematocrit, after 3, 6, 9 and 12 months of treatment as well as 3 months after withdrawal. Other paramenters after 52 weeks of exposure and 3 months of withdrawal.
- How many animals: each rat (haematocrit index).
- Parameters checked: hematocrit index of peripheral blood, haemoglobin, erythrocyte count, leukocyte count and composition, platelet count and reticulocyte percentage.

CLINICAL CHEMISTRY: Yes,
- Time schedule for collection of blood: every 3 months during treatment and 16 weeks after vanadyl withdrawal.
- Animals fasted: for 5 hr
- Parameters checked: aspartate aminotransferase, alanine aminotransferase and urea (for liver and kidney function).
- In addition, determination of non-fasting blood glucose (weekly in the first 4 weeks and then once upon every 2-4 weeks), fasting plasma glucose and insulin, plasma trigycerides and cholesterol throughout the exposure time (every 3 months).

URINALYSIS: No

NEUROBEHAVIOURAL EXAMINATION: No

IMMUNOLOGY: Not specified

OTHER:
- Determination of systolic blood pressure, pulse rate after 3, 6, 9 and 12 months of treatment as well as 3 months after withdrawal using tail-cuff method.
- Vanadium levels were determined in the 5-hr fasting plasma samples obtained at the end of the 52 wk-treatment and at weeks 6 and 12 after vanadyl withdrawal.
- V levels were determined in plasma, bone and some of the inner organs after 1 year exposure and after 16 weeks of vanadyl withdrawal

Sacrifice and pathology:

After treatment for 52 weeks most of the rats were terminated for morphological studies with an overdose of halothane followed by decaptation. 8 rats from the exposed groups and 3 rats from the control group were kept and observed for a further period of 16 weeks (without exposure).

GROSS PATHOLOGY: Yes,
- Brain, thymus, lung, heart, liver, spleen, pancreas, adrenal gland, kidney and testis were weighed.

HISTOPATHOLOGY: Yes,
- Brain, thymus, lung, heart, liver, spleen, pancreas, adrenal gland, kidney and testis were examined.

Other examinations:

no

Statistics:

All results were expressed as mean and standard error of the mean. The data were analyzed using repeated measure or one-way ANOVA, as appropriate, followed by the Newman-Keul’s test, if required. The values of haematological indices of the same rats before and after the withdrawal of vanadyl sulphate were compared using an unpaired Student’s t-test. The level of significance was set at P<0.05. The data from the morphological studies are presented as incidence (%) of the specific morphological abnormalities in each organ and were analyzed with the Chi-Square test. The data of plasma levels of AST; ALT and urea, of organ weight/body weight ratio, and of plasma and tissue concentrations of vanadium are shown as mean and standard error of the mean, and were analyzed with two-way or one-way analysis of variance, as appropriate, followed by the Newman-Keul’s test, if required.

Clinical signs:

not specified

Mortality:

mortality observed, non-treatment-related

Description (incidence):

- one animal died in the high dose group for unknown reasons after 18 weeks of treatment.

Body weight and weight changes:

effects observed, treatment-related

Description (incidence and severity):

- Body weight gain was slightly reduced in the low and mid dose groups indicated by an increasing number of deviations from control values from week 13 onwards reaching occasionally statistical significance.
- Body weight gain was markedly and statistically significantly, at most time points, reduced in the high dose group after week 4.
For details on body weight, please refer to the field "attached background material".

Food consumption and compound intake (if feeding study):

no effects observed

Food efficiency:

not specified

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

no effects observed

Ophthalmological findings:

not specified

Haematological findings:

no effects observed

Clinical biochemistry findings:

no effects observed

Urinalysis findings:

not specified

Behaviour (functional findings):

not specified

Immunological findings:

not specified

Organ weight findings including organ / body weight ratios:

no effects observed

Gross pathological findings:

no effects observed

Neuropathological findings:

no effects observed

Histopathological findings: non-neoplastic:

no effects observed

Histopathological findings: neoplastic:

not specified

Other effects:

not specified

Details on results:

Only data on non-diabetic rats are considered. According to the authors, apart from the body weight changes, intakes of VOSO4 and plasma V concentrations, there were no significant differences in all other parameters among the V treatment groups. Therefore, the pooled results of the 3 treatment groups were used.

FOOD CONSUMPTION
- Food intake was not changed by V exposure compared to the controls.

WATER CONSUMPTION AND COMPOUND INTAKE (if drinking water study)
- Water intake was not changed by V exposure compared to the controls.
- Average intakes of VOSO4 were reported to be 34, 54 and 90 mg/kg bw/day (corresponding to 0.16, 0.25 and 0.41 mmol/kg/day).

HAEMATOLOGY
- Haematocrit did not show any differences between treatment and control groups.
- No differences between treatment and control groups were observed with respect to the haematological indices.

CLINICAL CHEMISTRY
- Apart from transient fluctuations, data on clinical chemistry were not changed by the treatment.

ORGAN WEIGHTS
- Relative organ weights of brain, thymus, lung, heart, liver, spleen, pancreas, kidney, adrenal, testis did not differ from controls.

GROSS PATHOLOGY
- Gross pathology did not reveal marked differences between treated animals and their controls.

HISTOPATHOLOGY: NON-NEOPLASTIC
- Histopathology did not reveal marked differences between treated animals and their controls.
- Due to the small differences compared to controls, it is difficult to assess whether inflammatory focal cell infiltration of pancreas, interstitual cell hyperplasia in the testis and Leydig cell tumours seen in the groups after 1 year of treatment (1 or 2 animals each) as well as after cessation (1 animal each) was treatment-related. In contrast to the authors’ opinion, no clear recovery of liver and kidney lesions was observed after 16 weeks of cessation.

OTHER FINDINGS
- Systolic blood pressure, and pulse rate did not show any differences between treatment and control groups.
- At the end of the treatment period, plasma samples contained 0.18, 0.31 and 0.46 µg V/ml.
- At weeks 6 and 13 following withdrawal of V, no detectable amounts of V were found.
- Non-fasting blood glucose and fasting plasma glucose were not influenced by VOSO4 treatment, but fasting plasma insulin was reduced compared to the controls.
- VOSO4 treatment had no effect on triglycerides and cholesterol compared to the controls.
- A dose-related increase of V concentrations in various tissues was observed with a ranking bone>kidney testis>liver>plasma>pancreas>brain. After 16 weeks of cessation, small concentrations of V could be detected only in brain and kidney indicating some affinity of V to thes tissues but not in plasma and in the other organs investigated.

Dose descriptor:

NOAEL

Effect level:

750 mg/L drinking water

Based on:

test mat.

Sex:

male

Basis for effect level:

body weight and weight gain

Remarks on result:

other: 54±4.1 mg/kg bw/day (based on fluid intake)

Dose descriptor:

NOAEL

Effect level:

16.9 mg/kg bw/day (nominal)

Based on:

element

Sex:

male

Basis for effect level:

body weight and weight gain

Remarks on result:

other: based on fluid intake

Critical effects observed:

not specified

After cessation of VOSO4 treatment, no differences between controls and V treatment groups were observed with regards to the biological parameters apart from slightly increasing body weight gain in the treatment groups adapting to normal values of the control group.

Conclusions:

Four groups of non-diabetic male rats received different concentrations of vanadyl sulfate (VOSO4) in drinking water for 52 weeks. The low dose group received 500 mg VOSO4/L in water for 52 weeks. The mid dose group received 500 mg/L for 1 week followed by 750 mg VOSO4/L in water for 51 weeks. The high dose group received 500 and 750 mg/L for 1 week each followed by 1250 mg VOSO4/L in water for further 50 weeks. Three recovery animals of each group were followed for further 16 weeks after cessation of VOSO4 treatment.
Treatment of male rats with different dose levels of vanadyl sulfate in drinking water corresponding to 34, 54 and 90 mg/kg bw/day over 52 weeks did not indicate severe signs of systemic toxicity under the conditions of this study. Body weights were dose-dependently reduced in treatment groups compared to controls, occasionally reaching statistical significance in the low and mid dose groups and at most time points in the high dose group. At study termination (week 52) significant difference in body weight (>10%) compared to control animals was observed in high dose animals only. Body weights of low and mid dose animals were lower (<10%) but not significantly different and thus considered not biologically relevant.
Based on significant and biologically relevant effects on body weight in high dose animals, the mid dose level of 54 mg/kg bw/day represents a NOAEL.

Reference 4

Endpoint:

sub-chronic toxicity: oral

Type of information:

experimental study

Adequacy of study:

weight of evidence

Reliability:

3 (not reliable)

Rationale for reliability incl. deficiencies:

significant methodological deficiencies

Remarks:

The following deficiencies were noted: test substance insufficiently described, number of animals per group too low for appropriate statistical analysis, only male rats were used, individual body weight data not given, clinical signs were not recorded, complete haematology missing for 100 and 150 ppm group, haematology not performed for remaining groups, clinical biochemistry not conducted, organ weights, necropsy and histopathology not performed/recorded, FOB not performed, thyroid hormones not determined, ophthalmological examination not performed, historical control data not provided, the applied doses were changed during the study.

Qualifier:

no guideline followed

Principles of method if other than guideline:

Measurement of the cystine content of the hair of rats fed diets containing vanadium pentoxide was undertaken to determine if there were changes in cystine content indicative of abnormal metabolism.
Rats were given diets supplemented with vanadium at both high and low levels. In a separate experiment, methionine was also included in the diet of vanadium fed rats to determine whether this sulfur amino acid would counteract the effects of vanadium.

GLP compliance:

not specified

Limit test:

no

Species:

rat

Strain:

Wistar

Sex:

male

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Weight at study initiation: 200 to 350 gm (Details for Low Level Vanadium exposure group)
- Diet: Purina Dog Chow Checkers, ground in a Wiley Mill.

Route of administration:

oral: feed

Vehicle:

other: Purina Dog Chow Checkers

Details on oral exposure:

Low Level Vanadium exposure:
- One group received a stock diet of Purina Dog Chow Checkers.
- A second group received the stock diet, with 25 ppm of vanadium incorporated in the form of vanadium pentoxide.
- 50 ppm of vanadium was similarly included in the diet of a third group.
- After 35 days, the levels of dietary vanadium were raised to 100 and 150 ppm, respectively, in order to elicit more pronounced differences.

High Level Vanadium exposure:
- A similar study was performed on rats fed the vanadium pentoxide at levels of 500 and 1,000 ppm of vanadium.
- A fourth group of five rats which served as group-paired fed control for those receiving 500 ppm was added.

High Level Vanadium with added methionine:
- dl-methionine in amounts of 1.6 % was added to the diet of one group of rats whose diet further contained 500 ppm V as the pentoxide.
- Another similar group was fed a diet containing 500 ppm V only
- A third similar group served as group-paired fed controls for the latter.

Analytical verification of doses or concentrations:

yes

Details on analytical verification of doses or concentrations:

- The metallic composition of the stock and the vanadium-supplemented diets were determined by spectrographic analysis.
- Stock diet of the rats contained an average of 0.7 γ of vanadium per gram.

Duration of treatment / exposure:

- Low Level Vanadium exposure: 103 days
- High Level Vanadium exposure: 75 days
- High Level Vanadium with added methionine exposure: 63 days

Frequency of treatment:

Daily

Dose / conc.:

25 ppm

Remarks:

(nominal vanadium content in diet; first 35 days; low level vanadium exposure group)

Dose / conc.:

100 ppm

Remarks:

nominal vanadium content in diet (36 -103 days; low level vanadium exposure group)

Dose / conc.:

50 ppm

Remarks:

(nominal vanadium content in diet; first 35 days; low level vanadium exposure group)

Dose / conc.:

150 ppm

Remarks:

nominal vanadium content in diet (36 -103 days; low level vanadium exposure group)

Dose / conc.:

500 ppm

Remarks:

vanadium with or without methionine; nominal in diet (High Level Vanadium with added methionine exposure group)

Dose / conc.:

500 ppm

Remarks:

nominal vanadium content in diet (High Level Vanadium exposure group)

Dose / conc.:

1 000 ppm

Remarks:

nominal vanadium content in diet (High Level Vanadium exposure group)

No. of animals per sex per dose:

- Low Level Vanadium exposure: 5 male rats per dose plus one control group of 5 rats
- High Level Vanadium exposure: 5 male rats per dose plus one control group of 5 rats and one group of 5 rats as group-paired fed control for those receiving 500 ppm.
- High Level Vanadium with added methionine: 6 rats in the 500 ppm group, 6 rats in the control-fed group, and 7 rats at 500 ppm + 1.6% dl-methionine.

Control animals:

yes, concurrent vehicle

Details on study design:

No data

Positive control:

No data

Observations and examinations performed and frequency:

LOW LEVEL VANADIUM EXPOSURE:

CAGE SIDE OBSERVATIONS: No data

DETAILED CLINICAL OBSERVATIONS: No data

BODY WEIGHT GAIN: Yes

FOOD CONSUMPTION AND COMPOUND INTAKE (if feeding study):
- Food consumption for each animal determined and mean daily diet consumption calculated as g food/kg body weight/day: Yes
- Daily records of food consumption were kept throughout the duration of the experiment.

FOOD EFFICIENCY: No data

WATER CONSUMPTION: No data

OPHTHALMOSCOPIC EXAMINATION: No

HAEMATOLOGY:
- Red blood cell count and hemoglobin level were determined.

CLINICAL CHEMISTRY: No

URINALYSIS: No

NEUROBEHAVIOURAL EXAMINATION: No

IMMUNOLOGY: Not specified

OTHER:
- Just prior to the introduction of vanadium into the experimental diets, samples of hair were clipped from all the animals.
- Other hair samples were taken at 60 and 103 days (Low Level Vanadium) or 36 and 75 days (High Level Vanadium).
- The hair samples were obtained by first skirting off and discarding about 1 to 1.5 cm of the hair back from the tips. An electric clipper was then used to cut of the hair close to the skin over an area from the shoulder to the base of the tail on the back of the animal. Samples were defatted by placing in evaporating dishes containing benzene for 30 minutes, rinsing with fresh benzene, draining, and air drying (Sullivan, M.X.; Hess, W.C., and Howe, P. E.: A Comparison of the Wool and Skins of Full-Fed and Maintenance-Fed Lambs, J. Agric. Res. 61: 877, 1940).
- Hair samples were stored in a vacuum desiccator over Drierite for 24 hours or until analyzed.
- Samples were analyzed for cystine content.

Sacrifice and pathology:

GROSS PATHOLOGY: No data

ORGAN WEIGHT: liver-weight to body weight ratio (HIGH LEVEL VANADIUM WITH ADDED METHIONINE EXPOSURE)

HISTOPATHOLOGY: No data

Other examinations:

No data

Statistics:

Analysis of variance was used.

Clinical signs:

not specified

Mortality:

not specified

Body weight and weight changes:

no effects observed

Food consumption and compound intake (if feeding study):

no effects observed

Food efficiency:

not specified

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

not specified

Ophthalmological findings:

not specified

Haematological findings:

no effects observed

Clinical biochemistry findings:

not specified

Urinalysis findings:

not specified

Behaviour (functional findings):

not specified

Immunological findings:

not specified

Organ weight findings including organ / body weight ratios:

no effects observed

Gross pathological findings:

not specified

Neuropathological findings:

not specified

Histopathological findings: non-neoplastic:

not specified

Histopathological findings: neoplastic:

not specified

Other effects:

not specified

Details on results:

LOW LEVEL VANADIUM EXPOSURE:

BODY WEIGHT AND WEIGHT GAIN/FOOD CONSUMPTION AND COMPOUND INTAKE (if feeding study)
- Food intake of all groups (total food intake: control: 1,996 gm/rat; 100 ppm group: 2,130 gm/rat; 150 ppm group: 2,019 gm/rat) was nearly the same.
- The vanadium test groups gained more weight than did the controls (average body weight gain: control: 101 gm/rat; 100 ppm group: 156 gm/rat; 150 ppm group: 146 gm/rat); this was not statistically significant.

HAEMATOLOGY
- At 100 and 150 ppm, lower erythrocyte counts were obtained (at study termination: control: 7.7; 100 ppm group: 6.8; 150 ppm group: 6.3)
- Hemoglobin level were decreased (at study termination: control: 15.0; 100 ppm group: 14.5; 150 ppm group: 13.7); this was not statistically significant.

OTHER FINDINGS:
- The cystine content of the hair of the control group increased with time, whereas that of the rats fed the 100 ppm vanadium diet remained nearly constant
- At the 150 ppm level, a decrease in the average hair cystine values occurred.
- The hair cystine of all the individual rats in the control group increased, the smallest increase amounting to 1.15 %.
- Of three rats in the vanadium groups which showed higher hair cystine after, the greatest single increase (0.76%) occurred in those receiving the 100 ppm V.
- The decrease in cystine are considered significant on a relative basis, i.e., comparing the vanadium-fed groups with their corresponding controls.
- The average change in cysteine was statistically significant different from the 100 ppm group and the 150 ppm group (at the 0.5% level).

HIGH LEVEL VANADIUM EXPOSURE:

CLINICAL SIGNS AND MORTALITY
- One rat died in the third week.

BODY WEIGHT AND WEIGHT GAIN
- A decreased rate of body-weight gain occurred in both high level vanadium treated groups, compared with that of controls
- The decreased weight gain was greater in both vanadium groups than in the group-paired fed controls, indicating that food restriction was not the sole factor of growth impairment
- Average body weight gains: control: 125 gm/rat; 500 ppm group: 42 gm/rat; food control group: 90 gm/rat; 1000 ppm group: - 2 gm /rat; this was not statistically significant.

OTHER FINDINGS
- The cystine content of hair of rats fed the higher levels of vanadium, 500 and 1,000 ppm, was depressed in comparison with that of the controls
- Average change in cystine: control: 0.44 +/- 0.20 %, 500 ppm group: -0.19 +/- 0.48%; food control group: -0.26 +/- 0.25%; 1000 ppm group: -0.76 +/- 0.13%; statistically significant at the 5% level).
- This finding corresponded to that at the lower levels, with the difference that the percentage decrease in cystine at the 1,000 ppm level was greater (0.76%) and occurred sooner (75 days) than at the 150 ppm level which required 103 days to produce a 0.41 % average decrease in cystine content.
- The 0.76 % decrease in hair cystine content at the 1,000 ppm level of vanadium feeding is highly significant on an absolute basis, i.e., comparing final and original cystine percentages within the same group.

HIGH LEVEL VANADIUM WITH ADDED METHIONINE EXPOSURE:

BODY WEIGHT AND WEIGHT GAIN
- The weight gain per 100 gm of food consumed by the rats on the methionine-vanadium diets somewhat improved, as compared with those on vanadium diets (5.66 gm. vs. 6.62 gm, not statistically significant).

ORGAN WEIGHTS
- Methionine did not significantly affect either the growth or the relative liver weight in vanadium-fed animals.
- The average values for these suggest a definite tendency toward improvement as a result of the administration of methionine.
- The methionine supplement, however, did not significantly change the liver-weight to body weight rations.

OTHER FINDINGS
- The methionine supplement, however, did not significantly improve the growth of the hair.
- It was noted that the hair growth of the methionine-fed animals was markedly inhibited (10 -90 %).

COMBINED FINDINGS:
- Substantiating evidence for chemical changes found in the hair of vanadium-treated animals were structural changes, both gross and microscopic.
- The regrowth of hair was sparser, coarser, and stiffer compared with the more fleece-like hair of the controls.

Dose descriptor:

NOEL

Effect level:

4.58 mg/kg bw/day (nominal)

Based on:

element

Sex:

male

Basis for effect level:

body weight and weight gain

Dose descriptor:

NOEL

Effect level:

150 ppm

Based on:

element

Sex:

male

Basis for effect level:

body weight and weight gain

Critical effects observed:

not specified

Conversion of NOEL (ppm in diet) to NOEL (mg/kg bw /d):

 

NOEL:                                     150 ppm V

Duration:                                103 d

Food intake:                          2,019 g/rat

Body weight (kg bw/rat):     0.5 kg

V ingested reported:            236 mg V/rat/103 d= 2.29 mg V/d

NOEL_corrected 4.58 mg V/kg bw/d

Conclusions:

Male rats were treated with vanadium pentoxide for up to 103 days. The experimental setup was split into three phases: 1. animals were exposed to 0, 100 and 150 ppm V for 103 days; 2. animals were exposed to 0, 500 and 1000 ppm V and one group-paired fed control for the 500 ppm group for 75 days; 3. animals were exposed to 0 and 500 ppm V and an additional group was fed with a diet containing 500 ppm V and 1.6% methionine for 63 days.
In all three setups body weight gain and food intake as well as amount of ingested vanadium were recorded. In the first two setups of the study the cysteine content of hair was determined three times (1. setup: day 0, 60 and 103; 2. setup: day 0, 36 and 75). Additionally, in the first setup, red cell count and haemoglobin was determined before study initiation and at study termination. In the third setup liver to body weight ratio was recorded.
According to the author, hair cysteine content decreased dose-dependently in animals exposed to 100, 150, 500 and 1000 ppm vanadium. The food intake was not significantly affected in animals exposed to vanadium. However, animals exposed to 500 and 1000 ppm vanadium showed significantly decreased body weight gains after 75 days (500 and 1000 ppm V) and 63 days (500 ppm V). Animals exposed to 100 and 150 ppm vanadium for 103 days showed slightly decreased red cell counts and haemoglobin levels. No effects on liver to body weight ratios were observed in animals exposed to 500 ppm vanadium for 63 days.
Since the effects on red cell count and haemoglobin in animals exposed to 100 and 150 ppm for 103 days were mild and within historical control ranges of this rat strain and age, the effects were considered to be negligible. Further, cysteine content of rat hair was slightly reduced but the adversity of this effect remains questionable. Since body weight and food consumption in animals exposed to 100 and 150 ppm V were normal and no further adverse effects were observed, mildly decreased values of cysteine in hair is considered not adverse. Consequently, 150 ppm V (equivalent to 4.58 mg V/kg bw/day) represents the NOEL of this study, based on the reduced body weight gain in animals exposed to 500 and 1000 ppm vanadium.

Endpoint conclusion

Endpoint conclusion:

adverse effect observed

Dose descriptor:

LOAEL

Study duration:

chronic

Species:

rat

Quality of whole database:

Several supportive studies exist. A number of studies are available where vanadium compounds were administered; however, they have involved different experimental approaches and designs as well as different dose regimens, and endpoints. For oral exposure, effects are more limited and the different experimental approaches lead to a variety of endpoints measured.

Repeated dose toxicity: inhalation - systemic effects

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed

Repeated dose toxicity: inhalation - local effects

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed

Dose descriptor:

NOAEC

22 mg/m³

Study duration:

subacute

Species:

rat

Repeated dose toxicity: dermal - systemic effects

Link to relevant study records

Reference

Endpoint:

short-term repeated dose toxicity: dermal

Data waiving:

other justification

Justification for data waiving:

other:

Critical effects observed:

not specified

Endpoint conclusion

Endpoint conclusion:

no study available

Repeated dose toxicity: dermal - local effects

Link to relevant study records

Reference

Endpoint:

short-term repeated dose toxicity: dermal

Data waiving:

other justification

Justification for data waiving:

other:

Critical effects observed:

not specified

Endpoint conclusion

Endpoint conclusion:

no study available

Additional information

Initial comment on grouping and read across – oral

The endpoint repeated dose toxicity via the oral route of the vanadium category substance is not addressed by substance-specific information but rather by read-across of data available for soluble tri-, tetra- and pentavalent vanadium substances as well as for insoluble vanadium substances with zero valency, such as vanadium carbide.

All vanadium substances within the read-across concept represent inorganic substances, including salts or oxides and form the vanadium category. For the substances of the inorganic vanadium substances category, it is assumed that the in vivo bioavailability of the vanadium varies in a predictable manner, which is dependent on the in vitro bioaccessibility of the respective vanadium substance, i.e. all members of the category liberate vanadium ions in aqueous media at different rates, inter alia depending on the chemical structure. Thus, this concept is based on the chemistry / composition of all substances and on experimental studies for (i) water solubility and (ii) in-vitro bioaccessibility: assessment of the solubility and speciation of vanadium substances in five different artificial physiological fluids. Robust summaries for these studies are provided in each registration dossier and a detailed evaluation of the relevance, reliability and adequacy of each study is presented in the individual study records. Based on the in vitro bioaccessibility and the available bridging studies, two read-across groups are defined for the systemic effects following oral exposure, i.e. (i) soluble vanadium substances and (ii) poorly soluble vanadium substances. This endpoint summary addresses the hazard data for the soluble read-across group. Further details on the read-across concept are presented in the report attached to IUCLID section 13.

Therefore, the remaining text in this chapter is generic for all vanadium substances of the soluble read-across group and has not been adapted on a substance-specific basis.

 

Oral - animal data:

A number of studies are available reporting findings observed mainly in rats but also in mice orally exposed to soluble tetra- and pentavalent vanadium substances. However, the studies applied different experimental designs and were targeted at investigating specific endpoints. Thus, the different experimental approaches led to results on a variety of endpoints, which is why the weight of evidence approach is applied.

Based on the available data, the following adverse effects were identified: body weight reduction, haematological effects and reduction of antioxidants in tissues and body fluids. These effects are the most consistently reported among a number of investigators (Mountain et al. 1953, Dai et al. 1985, Roberts et al. 2015 and 2019, Zaporowska et al. 1993, Scibior et al. 2005, 2006, 2012).

 

A detailed review of the oral repeated dose toxicity studies in animals is presented below.

 

Mountain et al. 1953: Male rats were treated with vanadium pentoxide for up to 103 days. The experimental setup was split into three phases: 1. animals were exposed to 0, 100 and 150 ppm V for 103 days; 2. animals were exposed to 0, 500 and 1000 ppm V and one group-paired fed control for the 500 ppm group for 75 days; 3. animals were exposed to 0 and 500 ppm V and an additional group was fed with a diet containing 500 ppm V and 1.6% methionine for 63 days. In all three setups body weight gain and food intake as well as amount of ingested vanadium were recorded. In the first two setups of the study the cysteine content of hair was determined three times (1. setup: day 0, 60 and 103; 2. setup: day 0, 36 and 75). Additionally, in the first setup, red cell count and haemoglobin was determined before study initiation and at study termination. In the third setup liver to body weight ratio was recorded. According to the author, hair cysteine content decreased dose-dependently in animals exposed to 100, 150, 500 and 1000 ppm vanadium. The food intake was not significantly affected in animals exposed to vanadium. However, animals exposed to 500 and 1000 ppm vanadium showed significantly decreased body weight gains after 75 days (500 and 1000 ppm V) and 63 days (500 ppm V). Animals exposed to 100 and 150 ppm vanadium for 103 days showed slightly decreased red cell counts and haemoglobin levels. No effects on liver to body weight ratios were observed in animals exposed to 500 ppm vanadium for 63 days.  Since the effects on red cell count and haemoglobin in animals exposed to 100 and 150 ppm for 103 days were mild and within historical control ranges of this rat strain and age, the effects were considered to be negligible. Further, cysteine content of rat hair was slightly reduced but the adversity of this effect remains questionable. Since body weight and food consumption in animals exposed to 100 and 150 ppm V were normal and no further adverse effects were observed, mildly decreased values of cysteine in hair is considered not adverse. Consequently, 150 ppm V (equivalent to 4.58 mg V/kg bw/day) represents the NOEL of this study, based on the reduced body weight gain in animals exposed to 500 and 1000 ppm vanadium. This study was rated as RL 3 due to significant methodological deficiencies.

 

Dai et al. 1994: Four groups of non-diabetic male rats received different concentrations of vanadyl sulfate (VOSO4) in drinking water for 52 weeks. The low dose group received 500 mg VOSO4/L in water for 52 weeks. The mid dose group received 500 mg/L for 1 week followed by 750 mg VOSO4/L in water for 51 weeks. The high dose group received 500 and 750 mg/L for 1 week each followed by 1250 mg VOSO4/L in water for further 50 weeks. Three recovery animals of each group were followed for further 16 weeks after cessation of VOSO4 treatment. Treatment of male rats with different dose levels of vanadyl sulfate in drinking water corresponding to 34, 54 and 90 mg/kg bw/day over 52 weeks did not indicate severe signs of systemic toxicity under the conditions of this study. Body weights were dose-dependently reduced in treatment groups compared to controls, occasionally reaching statistical significance in the low and mid dose groups and at most time points in the high dose group. At study termination (week 52) significant difference in body weight (>10%) compared to control animals was observed in high dose animals only. Body weights of low and mid dose animals were lower (<10%) but not significantly different and thus considered not biologically relevant. Based on significant and biologically relevant effects on body weight in high dose animals, the mid dose level of 54 mg/kg bw/day represents a NOAEL. This study was rated as RL 2, comparable to guideline study with acceptable restrictions.

 

Domingo et al. 1985: Sodium metavanadate in drinking water was given to four groups, each consisting of 10 male Sprague-Dawley rats, at concentrations of 0, 5, 10 and 50 ppm. Liver and renal function parameters were determined in blood at the end of exposure period and organ weights were taken (liver, kidneys, heart, spleen, lung). Heart, liver, lungs, kidneys, spleen, stomach, small and large intestine were histopathologically examined in three rats of each group. Oral administration of NaVO3 via drinking water to groups of male rats over 3 months at concentrations of 0, 5, 10 and 50 ppm caused mild, dose-dependent lesions in kidneys, lungs and spleen with the highest incidence in the 50 ppm group, and increased plasma concentrations of protein, urea and uric acid in the high dose group. Thus, the highest dose level (7.57 mg/kg bw/d NaVO3) represents a clear LOAEL, and the mid dose level (1.51 mg/kg bw/d NaVO3) represents a NOAEL. This study was rated as RL 3 due to significant methodological deficiencies.

 

Zaporowska et al. 1993: Male and female Wistar rats were orally exposed to ammonium metavanadate (0, 10 or 50 mg V/L ) in drinking water for 4 weeks. Body weight gain, food and fluid uptake as well as various haematological and biochemical parameters were determined. Effects on palatability of vanadium containing fluids was observed at 50 mg V/L (significant in males but not in females). Mild and not dose-dependent effects were observed on body weight gain. Additionally, slight but significant effects on erythrocytes, reticulocytes and haemoglobin as well as L-ascorbic acid level in plasma and erythrocytes were observed in both dose groups. As no dose dependency was observed, mild effects on body weight gain were considered negligible. Significant effects on haematological parameters were considered to be mild and within historical control ranges of this rat strain and age (discussed below). Effects on L-ascorbic acid level in erythrocytes and plasma were also not considered relevant since the adversity of this effect is unclear (discussed below) and no further adverse effect was observed. Based on this, the NOEL of this study is considered to be 50 mg V/L drinking water, which is equivalent to 4.93 and 6.65 mg V/kg bw/day in males and females, respectively. This study is well reported, however, no guideline was followed and only limited parameters were analysed. Erythrocytes and haemoglobin and reticulocytes were slightly but significantly changed in both dose groups and sexes. These changes were not considered to be adverse as they were minor compared to control and within historical control data of this rat strain and age (e.g. Kort, M. et al. (2020) and Giknis, M. & Clifford, C.B. (2008)). Apart from the above-mentioned minor effects on some haematological parameters, the level of L-ascorbic acid was significantly decreased in plasma of low and high dose males but not in females. The same author reported one year later (please refer to Zaporowska 1994) that male and female rats exposed to doses of up to 0.30 mg V/ml showed clearly reduced L-ascorbic acid values in liver, kidney, adrenals and spleen. Thus, exposure to ammonium metavanadate seems to correlate with decreased L-ascorbic acid values in different tissues and body fluids. However, as no systemic toxicity or any other effect was observed, it remains unclear whether mild to moderate L-ascorbic acid depression is or results in any adverse effects. In a publication of Chan & Reade (1996) Wistar Shionogi rats, unable to synthesize L-ascorbic acid, were supplemented with different doses of L-ascorbic acid to determine the L-ascorbic acid requirements in Wistar rats. After 26 weeks, all animals survived and showed no clinical signs of scurvy. The average weekly body weight gain was normal. A severe L-ascorbic acid deficiency would include perinasal and peri-and intra-oral haemorrhage, joint or intramuscular haemorrhage, weakened or fractured hind limbs, delayed wound healing and a failure to thrive (Clemetson, 1989). As none of these clinical signs were observed in studies reported by Zaporowska (1993, 1994) or Chan & Reade (1996), it is assumed that this mild to moderate L-ascorbic acid depression observed in this study is not an adverse effect. It has been demonstrated that the toxicity of vanadium increases with its valency. Thus, compounds containing pentavalent V, such as ammonium metavanadate, are most poisonous. It is known that pentavalent V enters cells through anion channels, i.e. phosphate or sulfate channels. In cells, pentavalent V is reduced to divalent VO by some reducing compounds such as L-ascorbic acid and thiol-containing cysteine. Thus, the reported reduction of L-ascorbic acid in plasma and erythrocytes and also in several tissues (Zaporowska, 1994) is most likely the result of an enhanced consumption/reduction activity of this compound. However, in contrast to humans, rodents are able to synthesize L-ascorbic acid. Thus, it can be assumed that this reduction of L-ascorbic acid will induce re-synthesis and reduction of L-ascorbic acid is an adaptive but not an adverse effect. Apart from that, it is also noteworthy that humans are not able to synthesize L-ascorbic acid. Based on this, it is assumed that the protective reduction of pentavalent V to divalent VO is performed by another reducing substance and thus, it remains questionable whether a reduction of L-ascorbic acid in rats is relevant for humans. This study was rated as RL 3 due to significant methodological deficiencies.

 

 

The following studies reported similar effects on haematological parameters and body weight and also a few other signs of systemic toxicity, but were regarded as supportive, as several shortcomings in study design and reporting led to limited relevance in risk assessment.  

 

Scibior et al. (2012) and Scibior et al. (2018): A group of 16 male Wistar rats received sodium metavanadate (13.03 mg vanadium/kg bw/day) in drinking water for 12 weeks. The following parameters were investigated: clinical signs, mortality, physical appearance, motor behaviour, food/water consumption, haematology, clinical chemistry, and urinalysis (iron concentration only). Furthermore, atomic absorption measurements of vanadium, magnesium, iron, copper and zinc were conducted in selected biological material (liver, spleen, faeces, red blood cells, and urine). Lastly, femoral diaphysis were prepared and analysed for oxidative stress markers (superoxide dismutase, catalase, glutathione peroxidase, glutathione reductase, glutathione S-transferase, total glutathione, glutathione disulfide, lipid peroxidation) and L-ascorbic acid (also in the liver) as well as metal content (vanadium, magnesium, calcium, iron, copper and zinc).

 

Scibior et al. (2012): After the administration of 13.03 mg/kg bw/day of sodium metavanadate in drinking water some rats had gastrointestinal disturbances. Loose stool/diarrhoea was observed. No mortality was observed in the study. Furthermore, the treatment with sodium metavanadate led i) to a decrease in fluid and food intake and body weight gain, ii)  to the development of microcytic-hypochromic anaemia in the animals with excessive iron disposition in liver and spleen, unaltered plasma iron level and enhanced zinc concentration in red blood cells, characterized by a reduced haemoglobin level and haematocrit, unchanged counts of erythrocytes and reticulocytes, lowered total iron-binding capacity and elevated transferrin saturation, iii) disturbed Cu homeostasis, but iv) did not influence the leucocyte count and the plasma total antioxidant status.

 

Scibior et al. (2018): Vanadium alone did not significantly alter the thiobarbituric reactive substances and the activity of superoxide dismutase compared with the control but reduced slightly the glutathione reductase activity and the L-ascorbic acid level. It also markedly lowered the activity of catalase and glutathione peroxidase but elevated the activity of glutathione transferase and the hepatic L-ascorbic acid level to some degree.

Further, the vanadium concentration in the bone of the treatment group increased, whereas that of magnesium decreased compared with those in the control group. The total content of zinc and calcium did not change markedly in the treatment group compared to the control group.

In conclusion, the results provided evidence that the concentration of vanadium did not enhance the lipid peroxidation in rat bones. Vanadium administered disrupted the antioxidant defence mechanisms and homeostasis of some metals in bone tissue, which may consequently affect the balance in the activities of osteoblastic and osteoclastic cells.

 

Scibior & Zaporowska 2007: Outbred 2-month old male albino Wistar rats received for a period of 12 weeks vanadium metavanadate in drinking water at a concentration of 0.1 mg V/mL (equivalent to 8.63 mg V/kg bw/day). The control group received deionized drinking water. Food, fluid and water intakes were monitored daily and rats were weighed weekly. After 12 weeks, liver and kidney weights were recorded and the content of L-ascorbic acid in liver and kidney was analysed. Further, GSH and GSSG were determined in liver and kidney. The content of vanadium (V) in feed was analysed by ICP-AES. V, Fe, Zn and Cu were determined in liver and kidney by ICP-AES. Significant effects on GSH and GSSG level and GSH/GSSG ratio were observed in liver and kidney. Moreover, relative weights of liver and kidney were significantly increased. However, the slight effect on organ weights is considered to be small and thus an adaptive but not an adverse effect.  Thus, based on the adverse effects on GSH and GSSG level and GSH/GSSG ratio in liver and kidney, the LOAEL of this study is considered to be 8.63 mg V/kg bw/day. This study is well reported, however, no guideline was followed and only limited parameters were analysed. As no details on food and water intake were reported, it remains unclear if the palatability of vanadium containing drinking water was decreased. The author reported that GSH and GSSG level were slightly but significantly decreased in liver and kidney and vanadium levels in both organs were significantly increased. The same author reported a few years before that the levels of L-ascorbic acid were decreased in liver, spleen, kidney and adrenals as well as in plasma and erythrocytes (Zaporowska 1993, 1994). Both, L-ascorbic acid and GSH/GSSG (a disulfide derived from two glutathione molecules), are involved in detoxifying pentavalent vanadium compounds (as discussed in Zaporowska 1993, 1994). As shown in this study, vanadium was present in liver and kidney while in both organs levels of GSH and GSSG were reduced. Thus, the reported reduction of GSH and GSSG was most likley the result of an enhanced consumption/reduction activity of these compounds during detoxification of pentavalent vanadium. The fact that all cells are capable of synthesizing GSH and consumption of GSH initiates re-analysis of GSH most likely explains the slightly increased organ weights of liver and kidney. Thus, it is assumed that slightly affected organs weights were an adaptive but not an adverse effect.

 

Zaporowska et al. 1994: As reported in several other studies, effects on palatability of vanadium containing fluids were observed in high dose groups. Significant and dose-dependent decreases were observed in L-ascorbic acid levels in spleen, liver, adrenals and kidneys in males exposed to 0.15 and 0.30 mg V/mL and females exposed to 0.05, 0.15 and 0.3 mg V/mL. However, as the adversity of this effect remains questionable (discussed below) and no systemic toxicity was observed, this effect is not considered to be adverse and thus not taken into account for NOAEL setting. Based on this, 0.30 mg V/mL (equivalent to 22.06 mg V/kg bw/day in males and 26.62 mg V/kg bw/day in females) represents the NOEL. This study is well reported, however, no guideline was followed and only limited parameters were analysed. Significant and dose-dependent decreases were observed in L-ascorbic acid levels in spleen, liver, adrenals and kidneys in males exposed to 0.15 and 0.30 mg V/mL and females exposed to 0.05, 0.15 and 0.3 mg V/mL. Zaporwoska et al. (1993) reported already one year before that L-ascorbic acid levels were decreased in plasma and erythrocytes in rats exposed to ammonium metavanadate (see discussion above).

Scibior et al. 2005: Male Wistar rats were exposed to sodium metavanadate in drinking water (0.1 mg V/mL) for 6 weeks. Calculated average uptake of V was 8.35 ± 0.70 mg/kg/day. Food and fluid intake was reduced, which is however most likely a result of reduced palatability. Body weight gain was slightly affected (approx. 14 % reduction) but not significantly different compared to controls. An increase in erythrocyte count, a decrease in the mean corpuscular haemoglobin (MCH) and mean corpuscular haemoglobin conc. (MCHC) was observed in the treatment group. No statistically significant decrease in the number of leukocytes, haematocrit and haemoglobin was observed in test animals. Based on these results, 8.35 mg V/kg bw/day is considered to be the lowest observed adverse effect level (LOAEL).

 

Scibior et al. 2006: In this study, vanadium at the dose of 10.7 mg/kg bw/d consumed by rats with their drinking water for 6 weeks caused a significant decrease in food and fluid intakes, body weight gain, RBC count, Hb level, and MCV and MCH values, whereas no significant differences in WBC count were observed in these animals compared to the control group. Additionally, vanadium at the tested dose resulted in a significant decrease in L-ascorbic acid concentration in plasma and caused a significant increase in MDA concentration in RBC, whereas no significant differences in Total Antioxidant Status level in plasma of rats were shown compared to the controls. Moreover, vanadium treated rats had a higher vanadium concentration in the plasma than the control animals. Based on these results, the lowest observed adverse effects level (LOAEL) is considered to be 10.69 mg V/kg bw/day.

 

Susic et al. 2006: The study was conducted in male rats with sodium metavanadate in the diet for 24 weeks. Groups of rats received 0, 300 or 3000 ppm NaVO3 in the diet. Investigations: systolic blood pressure, heart rate and body weight, renal functions, haematological parameters, cardiac output, total peripheral resistance, determination of plasma V concentration and weight of the right ventricular wall of the heart. Sub-chronic dietary treatment of rats with NaVO3 over 24 weeks (300 or 3000 ppm) did not affect blood pressure, but induced an increase in total peripheral resistance and a decrease in cardiac output in both groups. Haematocrit was significantly increased and plasma, blood and extracellular fluid volumes decreased in the high dose group. Renal function parameters were not affected by treatment of rats with sodium vanadate.

 

Higashino et al. 1983: Female SD rats either on normal or high potassium intake were exposed to 5 or 25 ppm vanadium metavanadate for 2 weeks. Body weight was recorded weekly and clinical signs as well as food intake were recorded. At study termination blood was collected and organ weights were recorded. Urine was examined twice for endogenous creatine and Na and K fractional excretion. Additionally, the vanadium concentration in different organs were analysed and the plasma concentration of Na and K was determined. Further, the specific activity of Na-K-ATPase in different tissues was determined. Vanadium exposure had no influence on clinical signs and food and water intake and body weight gain was not affected either. Haematological parameters and urinalysis were not affected by vanadium. Despite extremely high tissue levels of vanadium, no effect of the element on the basal activity of Na-K-ATPase could be observed. As no adverse effects, except elevated vanadium levels in different tissues, were observed, the NOAEL of this study is 25 ppm (equivalent to 2129.5 ± 91.2 meq/kg bw/day, highest concentration of rats exposed to normal potassium intake, high potassium concentration/intake was not considered here).

 

Conclusion

Altogether, available studies on oral tetra- and pentavalent vanadium compound exposure report significant and biological relevant effects on body weight, indicating that vanadium compounds induce systemic toxicity.

In addition, mild but significant effects on haematological parameter were consistently reported. Effects included reduced haemoglobin, reduced haematocrit, reduced mean cell haemoglobin concentrations, reduced mean corpuscular haemoglobin and mean corpuscular volume as well as elevated reticulocyte counts. Erythrocytes were consistently decreased except in the study of Scibior et al. 2005, which reported a slight but significant increase of erythrocytes in sodium metavanadate treated animals. In contrast, white blood cells were consistently unaffected. Although most of the reported effects on haematological parameters were only mild and in most cases within historical control ranges of the respective rat strain and age, the almost consistent reduction of erythrocytes and haematocrit and increase in reticulocytes indicates that tetra- and pentavalent vanadium compounds induce anaemic effects by reducing erythrocytes, which in turn triggers an erythropoietic response.

The fact that evidence of haematological effects was also observed following 90-day inhalation exposure to vanadium pentoxide, in the absence of other remarkable systemic toxicity (NTP, 2002), increases the confidence in this being an appropriate critical effect for oral exposure from the available dataset. Additional support for the reliability of this endpoint comes from a study by Hogan (Comparative erythropoietic effects of three vanadium compounds, 2000), where haematological effects were demonstrated following IV injection of three different vanadium compounds each with a different valence state (vanadium chloride (V-III); vanadyl sulphate (V-IV); and sodium orthovanadate (V-V)).

In addition to the haematological effects, Scibior and Zaporowska reported in several studies with sodium metavanadate or ammonium metavanadate, that the level of L-ascorbic acid and glutathione in plasma and erythrocytes as well as in different tissues is markedly decreased. The adversity of this effect remains unclear, as no other adverse effects were observed. However, a possible explanation for this significant reduction may be, that vanadium, known to be present in several tissues and body fluids, is detoxified by L-ascorbic acid and glutathione. Thus, the reduction is most likely the result of an enhanced consumption of these substances. However, as humans are not capable to synthesize L-ascorbic acid, it is unknown whether or not this effect is relevant for humans.

 

National Toxicology Programme

It is to be noted, that the registrant is aware that the National Toxicology Programme (NTP) in the US nominated tetra- and pentavalent vanadium forms (sodium metavanadate, NaVO3, CAS 13718-26-8; and vanadium oxide sulphate, VOSO4, CAS 27774-13-6), i.e. species present in drinking water and dietary supplements in 2007 (http://ntp.niehs.nih.gov/). The NTP testing program foresees the conduct of animal studies via oral route (drinking water and feed) on VOSO4 & NaVO3 as follows:

- Genetic toxicology studies, i.e. the Salmonella gene mutation assays, with NaVO3 and VOSO4 - negative

-14 days dose-range finding experiments in Sprague-Dawley rats and B6C3F1/N mice (dose rats and mice: 0, 125, 250, 500, 1000, 2000 mg/L) – completed (Roberts et al. 2015)

- 90 days with Sprague-Dawley rats and B6C3F1/N mice (dose rats and mice: 0, 31.3, 62.5, 125, 250, or 500 ppm) – ongoing, first results presented below (Roberts et al. 2019)

- Perinatal dose-range finding study: gestation day 6 (GD 6) until postnatal day 42 (PND 42) with Harlan Sprague-Dawley rats - ongoing, first results presented below (Roberts et al. 2019)

- 28 days immunotoxicity study (dosed-water) with female B6C3F1/N mice (dose: 0, 31.3, 62.5, 125, 250, or 500 ppm) - ongoing

It can reasonably be anticipated that these studies will be of high quality and relevance, and thus will serve as a more robust basis than the current data base with all its shortcomings.

 

In the study by Roberts et al. 2015, vanadium oxide sulfate was administered in drinking water at concentrations of 83.8, 167.5, 335, 670, and 1340 mg/L (equivalent to 11.7, 23.2, 35.6, 54.5, and 77.2 mg/kg/day for males and 10.0, 16.9, 28.4, 40.7, and 56.0 mg/kg/day for females). No mortality was observed at any dose, no clinical signs were observed in the 83.8, 167.5, and 335 mg/L groups, whereas clinical abnormalities (thin appearance, hunched posture, and ruffled coats) were seen in male and female mice in the 670 and 1340 mg/L groups, which were related to the body weight effects. At the 83.8, 167.5, and 335 mg/L groups group mean body weight of males and females were comparable with the vehicle group. Body weight reductions were observed in the 670 and 1340 mg/L groups, which however were significant only in the 1340 mg/L groups at study termination. Decreases in feed consumption were observed at 1340 mg/L in males and females, and were consistent with body weight effects. Water consumption per kilogram body weight (g/kg/day) decreased with increasing vanadium oxide sulfate concentration for both male and female mice. Although test item intake (mg/kg/day) increased with increasing concentration in drinking water, the increase was not dose proportional due to decreased palatability. Lastly, numerous statistically significant organ weight alterations were evident, although the vast majority of these alterations were proportional to, or at least followed the same trend as, terminal body weights. Reductions in absolute thymus and thymus-to-body weight ratios were apparent for male and female mice in the 1340 mg/L group, and in females in the 670 mg/L group, possibly due to stress associated with substance exposure and reduced water intake.

Overall, a confounding variable in this study is the reduced water consumption, which appears to be a cause of decreased palatability. The reduced water consumption observed in this study was accompanied with reduced body weight at the highest dose levels.

Due to the limited study design typical for a dose-range finding experiment (detailed clinical observations, haematology, clinical chemistry, and histopathology missing; limited organ weight determination) and severe confounder of reduced water intake, the study is used as supplementary information only.

Sodium metavanadate exposure in drinking water resulted in survival of 100 % of males and females in the 0, 125 and 250 mg/L groups, 80 % of males in the 500 mg/L group, and all females in the 500 mg/L group (125, 250 and 500 mg/L are equivalent to 18.364, 36.065, and 58.216 mg/kg/day for males and 15.247, 26.446, and 38.455 mg/kg/day for females, respectively). Due to early deaths, clinical observations, reduced body weights, and decreases in water and food consumption, animals in the 1000 and 2000 mg/L groups (equivalent to 59.587 and 109.377 mg/kg/day for males and 48.750 and 46.736 mg/kg/day for females, respectively) were terminated early on Day 8 (males) and Day 9 (females). Clinical abnormalities were limited to males and females in the 500, 1000 and 2000 mg/L groups, including shallow or rapid breathing, ruffled coat, abnormal gait, hunched posture, lethargy and/or thin appearance.

At study termination, body weight reduction was apparent in males and females in the 500 mg/L group. Reductions in food mean consumption (g/day) were no longer apparent when food consumption was normalized to body weight (g/kg/day) suggesting food intake was proportional to body weight at the 500 mg/L concentration. Effects on water consumption were apparent for males and females in the 500 mg/L groups, but when normalized to body weight, these effects indicated water consumption was proportional to body weight for these groups. Mice in the 1000 and 2000 mg/L groups exhibited more pronounced toxicity (i.e., mortality, moribundity, and clinical abnormalities) related to body weight effects whereas 80 % of males and 100 % of females in the 500 mg/L group survived to study termination. Thus, palatability issues may have contributed to body weight reductions in the 1000 and 2000 mg/L groups.

Organ weight alterations were noted in the heart, lungs, and ovary (right and left) across all exposure concentrations, whereas reductions in thymus and liver weights were noted in the 500 mg/L group only.

Overall, a confounding variable in this study is the reduced water consumption, which might be due to palatability, and the potential dehydration of the animals. Some of the clinical signs can be attributed to dehydration. In addition, lower water consumption might have caused the body weight reduction observed at the 2000 mg/L dose level.

Due to the limited study design, short exposure duration of only 14 days, restricted examination of the animals (detailed clinical observations, haematology, clinical chemistry, and histopathology missing; limited organ weight determination), and lack of reporting the individual data, the study will not be used for hazard and risk assessment purposes but as supplementary information.

In the study by Roberts et al. (2019), time-mated F0 Hsd:Sprague-Dawley rats were exposed to sodium metavanadate and vanadyl sulfate at concentrations of 0, 31, 63, 125, 250, and 500 mg/L and 0, 21, 42, 84, 168, and 335 mg/L, respectively via drinking water on GD 6. Groups of male and female F1 animals were exposed during gestation, lactation and 13-weeks post-weaning (5 animals for biological sampling). Dams/pups exposed to sodium metavanadate at 250 and 500 mg/L in drinking water exhibited moribundity at birth, failure to thrive, and higher moribundity during lactation. Lower body weights were observed in dams during gestation and lactation, and in pups continuing until study termination 13 weeks post-weaning. Vanadyl sulfate, up to 335 mg/L in drinking water, was well tolerated in time-mated rats during gestation and lactation, and their pups during lactation and up to 13-weeks post weaning.

In study by Roberts et al. (2019), groups of 10 male and 10 female B6C3F1/N mice were exposed to sodium metavanadate and vanadyl sulfate at concentrations of 0, 31, 63, 125, 250, and 500 mg/L and 0, 21, 42, 84, 168, and 335 mg/L, respectively via drinking water for a duration of 13 weeks. Sodium metavanadate in drinking water resulted in lower body weights and water consumption at 500 mg/L in adult mice. Mice exposed to higher concentrations of sodium metavanadate also had lower thymus weight. Lastly, mice exposed to higher (125, 150, 500 mg/L) concentrations of sodium metavanadate exhibited increase in erythrocytes and reticulocytes and decreases haematocrit and haemoglobin. Vanadyl sulfate, up to 335 mg/L in drinking water, was well tolerated in adult mice.

The results of NTP studies by Roberts et al presented above are not yet fully accessible or published, since the review process is still ongoing. The results presented above are based on a 2019 congress abstract/poster. A complete robust study summary will be provided in the dossier upon availability of the data.

Inhalation:

The most informative study is the standard NTP chronic inhalation study (NTP 2002) using V2O5. In this investigation, there was a statistical increase in lung tumours in mice of both sexes, but not in rats (Starr, 2012). In mice, survival rates of male mice exposed to 4 mg/m3 was less than that of chamber controls, and mean body weights of male mice exposed to 4 mg/m3 and all exposed groups of female mice were generally less than those of the chamber controls throughout the study. As in the 3-month studies, the respiratory tract was the primary site of toxicity. Under the conditions of this 2-year inhalation study there was clear evidence of carcinogenic activity of vanadium pentoxide in male and female B6C3F1 mice based on increased incidences of alveolar/bronchiolar neoplasms. Exposure to vanadium pentoxide caused a spectrum of non-neoplastic lesions in the respiratory tract (nose, larynx, and lung) including alveolar and bronchiolar epithelial hyperplasia, inflammation, fibrosis, and alveolar histiocytosis of the lung in male and female mice. Hyperplasia of the bronchial lymph node occurred in female mice. The lowest concentration tested (1 mg/m3) represents a LOAEC for local effects in the respiratory tract.

 

Pulmonary reactivity was also investigated in a subchronic inhalation study in cynomolgus monkeys (duration 6 months) with divanadium pentaoxide. The results showed a concentration-dependent impairment in pulmonary function, characterized by airway obstructive changes (pre-exposure challenges) accompanied by a significant influx of inflammatory cells recovered from the lung by bronchoalveolar lavage. Subchronic V2O5 inhalation did not produce an increase in V2O5 reactivity, and cytological, and immunological results indicate the absence of allergic response.

 

However, local effects on the respiratory tract are not considered relevant for vanadium dioxide for the following reasons:

Severe irritant properties of V₂O₅have been identified for eye (cat 1) and in lungs, and the redox potential of V2O5 as well as the sharp decline on pH in contact with aqueous media is hypothesised to either mediate this mechanism or at least propagate this mechanism. In contrast, there is no indication whatsoever of any potential for irritation of the respiratory tract for V₂O₃and VO₂. With regard to substance-specific properties assumed to predominantly account for an irritation potential, V2O3 and VO2 are different from V₂O₅as follows:

- A low dissolution of V₂O₃was observed in artificial lysosomal fluid (11.9% after 2h; 15.7% after 24h) and lung fluid (4.7% after 2h; 5.6% after 24h)while pentavalent substances (V₂O₅or NaVO₃) dissolved completely within 2 h. Thus, the bioavailability and reactivity of V₂O₃in tissues of the respiratory tract is assumed to be far less than that of V₂O₅. Based on similar water solubility and transformation/dissolution characteristics, a similar low bioavailability in the lung is assumed for VO2.

- V₂O₃and VO₂upon contact with water do not cause such significant pH decrease as is the case for V₂O₅, thus indicating a lack of acidifying properties in aqueous media and any potential for tissue injury associated therewith of V₂O₃and VO₂.

- V₂O₃and VO₂are completely void of oxidising properties and the potential for oxidative injury.

- While VO₂lacks any irritation potential to skin and eye in vivo, some very mild but reversible effects have been observed in vivo in the eye after exposure to V₂O₃.

-V₂O₃and VO₂are not acutely toxic or harmful via inhalation whereas V2O5 is.

Regarding the potential for respiratory irritation, a comprehensive histopathological evaluation of lung tissue was performed within 14-d inhalation studies conducted both with V₂O₃and V₂O₅. Severe lung effects including hyperplasia in alveolar and bronchial epithelia, inflammation or fibrosis could not be observed at exposure levels up to 250 mg/m3 with V2O3, whereas these effects are reported as severe for all animals exposed to V₂O₅already at a level of 2 mg/m3. In conclusion, the onset of marked irritation effects with V2O5 occurs at exposure levels approx. 100-fold lower than with V2O3; on the other hand, given the low solubility and the high exposures, the onset of overload phenomena cannot be completely excluded for V2O3. Based on similar substance-specific properties (as outlined above) this conclusion is read-across to VO2. Thus, it is assumed that vanadium dioxide does not cause respiratory tract irritation.

 

No carcinogenicity, no pneumoconiosis and no other signs indicative of allergic inflammation have been reported for workers manufacturing vanadium dioxide.Therefore, the local respiratory effects of V2O5 are not relevant for read-across to vanadium dioxide.

 

The registrant is aware that the National Toxicology Programme (NTP) in the US nominated tetra- and pentavalent vanadium forms(sodium metavanadate, NaVO₃, CAS # 13718-26-8; and vanadium oxide sulphate, VOSO₄, CAS # 27774-13-6), i.e. species present in drinking water and dietary supplements in 2007 (http://ntp.niehs.nih.gov/). A comprehensive characterisation via the oral route of exposure of

(i) chronic toxicity,

(ii) carcinogenicity, and 

(iii) multi-generation reproductive toxicity

is planned.

 

The NTP testing program began with sub-chronic drinking water and feed studies on VOSO₄& NaVO₃as follows:

- Genetic toxicology studies, i.e. the Salmonella gene mutation assays, with NaVO3 and VOSO4 - negative

-14 days with Harlan Sprague-Dawley rats and B6C3F1/N mice (dose: R&M: 0, 125, 250, 500, 1000, 2000 mg/L) - already completed

- 90days with Harlan Sprague-Dawley rats and B6C3F1/N mice (dose: R&M:: 0, 31.3, 62.5, 125, 250, or 500 ppm) - ongoing

- Perinatal dose-range finding study: gestation day 6 (GD 6) until postnatal day 42 (PND 42) with Harlan Sprague-Dawley rats - ongoing

- 28days immunotoxicity study (dosed-water) with female B6C3F1/N mice (dose:0, 31.3, 62.5, 125, 250, or 500 ppm) - ongoing

 

It can reasonably be anticipated that these studies will be of high quality and relevance, and thus will serve as a more robust basis than the current data base with all its shortcomings.In addition, repeated-dose inhalation toxicity studies (14, 28, and 90 days) with various vanadium substances are planned within the Vanadium Safety Readiness Safety Program. These studies will address issues for which to date equivocal or no data at all exist.Further information on these studies can be found in the attachments below.Only upon availability of the results from these studies, it will be possible to render a more meaningful decision on whether or not testing for repeated-dose toxicity is required. Therefore for the time being this data requirement should be waived in consideration of animal welfare.

 

Justification for classification or non-classification

The currently available and reliable toxicity data on vanadium substances does not justify classification of vanadium dioxide for specific target organ toxicity - repeated exposure.