Registration Dossier

Administrative data

Description of key information

NOAEL for systemic toxicity, oral, from a sub-chronic study in rats: 17 mg Mo/kg bw/day.
NOAEC for systemic toxicity, inhalation, from a sub-chronic study in rats and mice: 66.7 mg Mo/m³.

Key value for chemical safety assessment

Additional information

As discussed in the dossier section on toxicokinetics, read-across between the substances registered by the molybdenum consortium is considered feasible without restriction, since the molybdate ion is the only relevant molybdenum species released from any of the molybdenum substances under physiological conditions. Sodium/ammonium/calcium/iron moieties are not considered to represent any toxicological concern based on their ubiquitous physiological presence in biota and/or their essential role in human physiology.

 

 

Key study on oral repeated dose toxicity:

 

In a 90-day repeated dose toxicity study which also included a 60 day recovery phase, disodium molybdate was administered to male and female rats at doses of 5, 17 or 60 mg/kg bw/day of Mo (administered as sodium molybdate dihydrate via feed). This study was conducted according to OECD Guideline 408, with additional consideration of parameters related to reproductive toxicity, as specified in OECD 416. In addition to the standard examination parameters, the following examinations were conducted to assess any adverse effects on sexual function and fertility: vaginal cytology, oestrous cycle, sperm parameters (count, motility and morphology, testicular spermatid counts).

Reduced bodyweight gains were observed only in the 60 mg Mo/kg bw/day dose group. The effect was more pronounced in males, which was partly due to a slightly reduced food intake and partly due to reduced food conversion efficiency.

During the recovery phase food consumption in the 60 mg/kg bw/day males and females returned to a value comparable to the control animals. Light microscopic evaluation of control and 60 mg Mo/kg bw/day animals showed test item-related findings in the kidneys (slight diffuse hyperplasia of the proximal tubules) of two 60 mg Mo/kg bw/day females. No such findings were reported for the animals after the 60-day recovery phase.

Compared to controls, serum copper levels, and liver and kidney copper concentrations, were significantly increased in both males and females in the group given the highest dose of 60 mg Mo/kg bw/day. However, these alterations in copper levels were not associated with any signs of toxicity, and the levels of copper in the liver were well below those associated with liver toxicity in other studies (Hébert 1993; Stern 2010).

 

 

There were no test substance related changes in male or female reproductive tissues (testes, epididymis, prostate, seminal vesicles, ovaries, uterus or vagina) at any dose level. There were also no test substance related effects on vaginal cytology and oestrous cycles during weeks 7-9 of the dosing phase (i.e., the period during which vaginal cytology and oestrous cycles were evaluated). No test item related changes in organ weight of testes or secondary sex organs and no effects on spermatid or sperm counts, motility or morphology were observed. All other recorded microscopic findings were considered incidental and unrelated to administration of disodium molybdate dihydrate. They occurred at similar incidences in the control and test substance treated groups or they were sporadic with no relationship to dose.

The NOAEL was determined to be 17 mg Mo/kg bw/day based on the effects on body weights and kidneys seen at 60 mg Mo/kg bw/day. The NOAEL for effects on reproductive organs, sperm and oestrous cycle is 60mg Mo/kg bw/day.

 

Prior to this 90-day study, a 28-day study was conducted as a dose-range-finding- and toxicokinetic study for the subsequent repeated-dose 90-day study. To investigate possible differences in uptake and to select an appropriate type of administration for the 90-day study (via gavage or via food), both types were used in separate groups in this 28-day dose-range-finding study. For technical reasons (IUCLID), there are two separate study records for administration via gavage and via food. Details on these 28-day-studies can be found in the technical dossier and in the corresponding tables in the CSR.

 

 

Key studies on repeated dose inhalation toxicity:

 

A 13-week inhalation toxicity study with molybdenum trioxide in rats and mice (NTP, 1997) was conducted in accordance with OECD test guideline 413 (1981) and also in compliance with FDA GLP Regulations, 21 CFR, Part 58. The study results are well-documented; historical control data are also included. The study represents a highly reliable study without restrictions. Thus, it is considered as the key study for risk assessment purposes of systemic effects of molybdenum compounds after inhalation exposure. 10 male + 10 female rats or mice per group were exposed in chambers to 0, 1, 3, 10, 30, 100 mg MoO3/m³ for 6.5 hours per day, 5 days per week for 13 weeks. The test substance is characterised as follows: MoO3, purity: ca. 99%, particle size: MMAD (µm) ± GSD in the range from 1.33 ± 1.93 to 1.60 ± 1.83.

Finding (rats): At all exposure concentrations, no treatment-related effects on mortality, clinical signs, final mean body weights, organ weights, haematology or clinical chemistry parameters, sperm counts or motility and liver copper concentrations were observed at all concentrations. No treatment-related gross or microscopic lesions were observed. Thus, the concentration of 100 mg MoO3/m³ (corresponding to 66.7 mg Mo/m³) represents a true NOAEC in this 13-week inhalation study on rats, since no adverse effects were seen up to and including the highest concentration tested.

Findings (mice): There were no adverse treatment-related effects on mortality, clinical signs, final mean body weights, organ weights, haematology or clinical chemistry parameters, and epididymal weights, sperm counts, or motility were observed at all concentrations. Also, no treatment-related gross or microscopic lesions were observed. There were significant increases in liver copper concentrations in female mice exposed to 30 mg/m3 and in male mice exposed to 100 mg/m³ compared to those of the control groups, which were not considered adverse due to a lack of any toxicological correlate. Thus, similar to rats, the 13-week inhalation study on mice results in a NOAEC of 100 mg MoO3/m³ (corresponding to 66.7 mg Mo/m³).

 

The NTP further conducted 2-year inhalation studies in rats and mice at 0, 10, 30 and 100 mg MoO3/m³ .In addition to their focus on local effects in the lung, a comprehensive set of systemic end points were studied and included body weight changes, reproductive parameters, and full histological evaluation of a wide range of tissues including the reproductive organs. The substance-specific local effects observed following inhalation of MoO3are discussed in the dossier section on carcinogenicity. Regarding systemic effects, despite the longer exposure duration, no adverse systemic effects were observed in the 2 year studies in rats and mice and both the 13-week and 2-year inhalation studies resulted in identical NOAECs for systemic toxicity of 100 mg MoO3/m³.

 

 

 

Lack of relevance of repeated dose toxicity following dermal exposure:

 

Firstly, the dermal absorption of molybdate anions has been shown experimentally to be low to negligible. Secondly, molybdenum substances are void of toxicity upon acute exposure via the dermal route, and are of very low systemic toxicity upon repeated oral and/or inhalation exposure. For this reason, toxic effects of molybdenum substances after repeated exposure via the dermal route are not considered to be of concern and therefore the conduct of toxicity studies in laboratory animals via repeated dermal exposure is not considered to be required.

 

 

Other data:

 

The registrant, via the Molybdenum Consortium and the International Molybdenum Association, has conducted an extensive literature/data search and evaluation programme on animal and human repeated dose toxicity data of molybdenum substances. All data sources were assessed by expert toxicologists for quality and reliability, as well as relevance for regulatory risk assessment under REACH. A full evaluation report is attached to the technical dossier. In conclusion, out of published data, only the inhalation toxicity experiments conducted with molybdenum trioxide in the context of the NTP (1997) toxicology and carcinogenesis studies in rats and mice are considered to be sufficiently robust and reliable for regulatory risk assessment of molybdenum compounds under REACH. All other references evaluated in this process lacked relevance because of severe shortcomings in the applied methodology and/or quality of reporting/documentation, thereby rendering them unsuitable for regulatory purposes. In adaption of the scoring scheme developed by Klimisch et al (1997), reliability scores of either “3 = not reliable” or “4 = not assignable” were assigned to all of these references except for the NTP study. For enhanced IUCLID-readability only key disregarded studies/references are included in the technical dossier. Other disregarded studies are viewable in attached evaluation report.

 

Human study reported by Kovalskiy et al. (1961):

A detailed review and critique of this publication is attached to the corresponding endpoint record in the technical dossier. In summary, based on a variety of methodological and reporting deficiencies, the publication by Kovalskiy et al. (1961) is considered an unreliable reference for the assessment of potential adverse effects of molybdenum on human health.

 

Executive summary of attached critique on publication by Kovalskiy et al. (1961):

The publication by Kovalskiy et al. (1961) attempts to establish a correlation between molybdenum content in soil, uptake via plants and thus via food, and corresponding human molybdenum blood levels. In turn, higher molybdenum blood levels are then thought to be correlated with an increase in serum xanthine activity, ultimately postulated to elicit gout by raising uric acid levels. This critique addresses in detail the question whether the mechanism postulated by Kovalskiy is scientifically plausible: in other words, is it really reasonable to assume that an increase in molybdenum blood levels can increase uric acid levels by an increase in serum xanthine oxidase activity?

A subchapter of this document describes the function and properties of xanthine oxidase. Based on this, there is no reason to assume that increased blood XO is correlated with gout-like symptoms. Based on the date of publication, the degree of sophistication and precision of the xanthine oxidase enzyme assay as well as the method for analysing uric acid in blood used by Kovalskiy falls short of current standards, and therefore raises sincere doubts as to the reliability of the measured data.

As far as analysis of Cu and Mo in blood, soil and plant matter is concerned, in retrospective it is not possible whatsoever to assess the validity of the analytical results presented because of missing auxiliary information and in particular the complete absence of any method validation data. The absence of such information represents a major deficiency in conduct and reporting.

We also question whether the increased molybdenum intakes postulated by Kovalskiy would consistently affect human molybdenum blood levels, and whether the intakes described would yield blood levels outside the human homeostatic control range. A comparison of what Kovalskiy himself designates as “controls” with current blood reference ranges for molybdenum and copper reveals that already his control levels by far exceed current reference ranges, thus leaving his further conclusions rather questionable.

Finally, we have undertaken a detailed review of the mathematical and statistical methodology applied. The outcome of this was that the publication is flawed by an inappropriate diagnosis of gout and comparison with statistical data, as well as an imprecise description of the study population and wrong conclusions based on different sub-groups. Any conclusions drawn on a combination of statistical analysis results in different sub-groups are therefore not appropriate, and a correlation of molybdenum in blood levels with xanthine oxidase activity cannot be considered as being statistically established.

Further, when taking into account an alpha adjustment, healthy Ankavana persons do not have significantly different xanthine oxidase activities compared to the control group, which implies that the higher molybdenum in blood level in “Ankavana province” does not lead to significantly higher xanthine oxidase activity in all persons but merely in “gout-sick” persons.

In conclusion, based on a variety of methodological and reporting deficiencies, the publication by Kovalskiy et al. (1961) is considered an unreliable reference for the assessment of potential adverse effects of molybdenum on human health.

 

 

References:

 

Hébert CD et al. (1993) Subchronic toxicity of cupric sulfate administered in drinking water and feed to rats and mice. Fundamental and Applied Toxicology, 21:461–475.

 

Stern, B.R. (2010). Essentiality and toxicity in copper risk assessment: Overview, update and regulatory considerations. . J. Toxicol. Environ. Health A 73:114-127.

 

Justification for classification or non-classification

The available, reliable toxicity data on molybdenum substances does not justify classification for specific target organ toxicity - repeated exposure.