2-ethylhexanoic acid, molybdenum salt

Repeated dose toxicity: inhalation

Administrative data

Endpoint:

sub-chronic toxicity: inhalation

Type of information:

experimental study

Adequacy of study:

key study

Study period:

1983

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

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

Data source

Reference

Reference Type:

publication

Title:

Unnamed

Year:

1997

Materials and methods

Test guideline

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 413 (Subchronic Inhalation Toxicity: 90-Day Study)

Deviations:

no

GLP compliance:

yes

Limit test:

no

Test material

Test material information

Constituent 1

Reference substance name:

Molybdenum trioxide

EC Number:

215-204-7

EC Name:

Molybdenum trioxide

Cas Number:

1313-27-5

Molecular formula:

MoO3

IUPAC Name:

molybdenum trioxide

Details on test material:

- Name of test material (as cited in study report): Molybdenum trioxide
- Physical state: Powder
- particle size: the test material was micronised to an MMAD of approx. 1.5 micrometers (analysed on a weekly basis, see details in the full study report)
- Analytical purity: approx. 99 %
- Impurities (identity and concentrations): Cadmium ≤ 100 ppm; potassium 2400 ppm; and silicon 180 ppm)
- Stability: The stability of the bulk chemical was monitored periodically by the study laboratories using atomic absorption spectroscopy and gravimetric analysis. No degradation of the bulk chemical was observed.
- Storage condition of test material: stored under refrigeration when not in use andallowed to warm to room temperature overnight prior to use
- Karl-Fischer water analysis: 0.03 % +/- 0.02 % water
No further information on the test material was stated.

Test animals

Species:

mouse

Strain:

B6C3F1

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Age at study initiation: 6 weeks
- Housing: Individually in stainless steel mesh cages, changed twice weekly; No bedding/cageboard; Chamber filters: HEPA (intake) and charcoal and HEPA (exhaust); Stainless steel racks
- Diet (ad libitum except during exposure): NIH-07 open formula, pellets (Zeigler Brothers, Inc., Gardners, PA)
- Water (ad libitum): Tap water ( City of Vienna water supply)
- Quarantine period: 2 weeks before study; Before initiation of the studies, two male and two female mice were randomly selected for parasite evaluation and gross observation for evidence of disease.

ENVIRONMENTAL CONDITIONS
- Temperature: 20 °C - 31 °C
- Relative humidity: 55 % - 95 %
- Chamber air: 200 L/minute
- Photoperiod (hrs dark / hrs light): 12/12
No further information on the test animals was stated.

Administration / exposure

Route of administration:

inhalation: dust

Type of inhalation exposure:

whole body

Vehicle:

air

Remarks on MMAD:

MMAD / GSD: MMAD of approx. 1.5 micrometers (analysed on a weekly basis, see details in the full study report)

Details on inhalation exposure:

GENERATION OF TEST ATMOSPHERE / CHAMBER DESCRIPTION
Molybdenum trioxide was generated by Wright dust-feed mechanisms at gear ratios appropriate for each target concentration on top of approximately 1-L elutriators which opened into the top of each chamber. The airborne dust was swept into the chamber by compressor air at 30 psi and 200 L/minute. Chamber air pressure was negative with respect to that of the room.

TEST ATMOSPHERE
Aerosol concentration monitoring:
Gravimetric samples were obtained during exposure periods from closed-face Gelman DM-450 Metricel filters in each exposure chamber two to six times per day. Samples were analysed for molybdenum content by atomic absorption. A real-time aerosol monitor (RAM) (Model RAM-1; GCA Corp., Bedford, MA) was used to monitor chambers in real time during the exposure periods. Readings were recorded approximately hourly for each chamber and were used to make adjustments to dust generating systems.
Chamber atmosphere characterization:
Particle size distribution in each chamber was determined weekly for 6 or 7 weeks then again in week 11 or 12 during the 13 week study using an Anderson 8-stage cascade impactor with an 11-micron preseparator. An estimation was made of the mass median areodynamic particle diameter and the geometric standard deviation of each set of samples.
For the 13-week study, the time required to achieve 90 % of target concentration at the start of exposure (T90) was 23 minutes. The time required for the concentration to decay to 10 % of target at the end of exposure (T10) was 23 minutes.
No further details on inhalation exposure was stated.

Analytical verification of doses or concentrations:

yes

Details on analytical verification of doses or concentrations:

see "Details on inhalation exposure " for analytical verification of concentrations.
The means of concentration in all chambers during the 13-week studies were within 10 % of the target concentration.

Duration of treatment / exposure:

13 weeks

Frequency of treatment:

6.5 hours per day, 5 days per week

Doses / concentrations

Doses / concentrations 1

Dose / conc.:

0 mg/m³ air (nominal)

Doses / concentrations 2

Dose / conc.:

1 mg/m³ air (nominal)

Doses / concentrations 3

Dose / conc.:

3 mg/m³ air (nominal)

Doses / concentrations 4

Dose / conc.:

10 mg/m³ air (nominal)

Doses / concentrations 5

Dose / conc.:

30 mg/m³ air (nominal)

Doses / concentrations 6

Dose / conc.:

100 mg/m³ air (nominal)

No. of animals per sex per dose:

10 males / 10 females

Control animals:

yes

Details on study design:

- Dose selection rationale: The doses were selected based on a 14 day inhalation study with mice. Exposure of mice to molybdenum trioxide for 14 days at concentrations of 0, 3, 10, 30, 100, or 300 mg/m^3 had no effect on survival or clinical findings. However, final mean body weights of male and female mice exposed to 300 mg/m^3 were significantly lower than those of the control group. Because of the body weight effects in the 14-day study, mice were expsoed to 0, 1, 3, 10, 30, or 100 mg/m^3 molybdenum trioxide during the 13- week study.
No further details on study design was stated.

Positive control:

No data

Examinations

Observations and examinations performed and frequency:

CAGE SIDE OBSERVATIONS: Yes
- Time schedule: Weekly

DETAILED CLINICAL OBSERVATIONS: No data

BODY WEIGHT: Yes
- Time schedule for examinations: The animals were weighed initally, weekly, and at the end of the studies.

FOOD CONSUMPTION:
- Food consumption for each animal determined and mean daily diet consumption calculated as g food/kg body weight/day: No data

FOOD EFFICIENCY:
- 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: No data

WATER CONSUMPTION: No data

OPHTHALMOSCOPIC EXAMINATION: No data

HAEMATOLOGY: No data

CLINICAL CHEMISTRY: No data

URINALYSIS: No data

NEUROBEHAVIOURAL EXAMINATION: No data

- OTHER: Sperm motility
At terminal sacrifice, sperm samples were collected from 0, 10, 30, and 100 mg/m^3 mice. The parameters evaluated included:Spermatogenesis, sperm count and motility. For sperm analyses , the left epididymis and testis were isolated and weighed. The tail of the epididymis (cauda epididymis) was then removed from the epididymal body (corpus epididymis) and weighed.

Liver copper analysis:
All mice were evaluated for liver copper burden. Liver tissue was prepared by wet digestion with nitric and perchloric acids for copper analysis by atmomic absorption spectroscopy.

No further information on observations and examinationsperformed and frequency were stated.

Sacrifice and pathology:

A necropsy was performed on all animals. The brain, heart, right kidney, liver, lung, right testis, and thymus were weighed. A complete histopathologic examination was performed on 0 and 100 mg/m^3 mice. In addition to gross lesions and tissue masses, the tissue examined included: Adrenal gland, brain, esophagus, eye, femorotibial joint, gallbladder, heart, large intestine (cecum, colon, rectum), small intestine, kidney, larynx, liver, lung, lymph nodes, (mandibular, mediastinal, peribronchial), mammary gland, nose ( all animals in all exposure groups), ovary, pancreas, parathyroid gland, pituitary gland, prostate gland, salivary gland, seminal vesicle, spinal cord, spleen, sternum, stomach, testis and epididymis, thymus, thyroid gland, trachea, urinary bladder, uterus, and vagina.

Statistics:

The probability of survival was estimated by the product-limit procedure of Kaplan and Meier (1958) and is presented in the form of graphs. Statistical analyses for possible dose-related effects on survival used Cox’s (1972) method for testing two groups for equality and Tarone’s (1975) life table test to identify dose-related trends. All reported P values for the survival analyses are two sided.
Analysis of continuous variables:
Two approaches were employed to assess the significance of pair wise comparisons between exposed and control groups in the analysis of continuous variables. Organ and body weight data, which have approximately normal distributions, were analyzed using the parametric multiple comparison procedures of Dunnett (1955) and Williams (1971, 1972). Clinical chemistry, hematology, blood ,olybdenum levels, spermatid evaluations, liver copper levels, and bone density and curvature, which have typically skewed distributions, were analyzed using the nonparametric multiple comparison methods of Dunn (1964) and Shirley (1977). Jonckheere’s test (Jonckheere, 1954) was used to assess the significance of the dose-related trends and to determine whether a trend sensitive test (Williams’ or Shirley’s test) was more appropriate for pair wise comparisons than a test that does not assume a monotonic dose-related trend (Dunnett’s or Dunn’s test). Average severity values were analyzed for significance using the Mann- Whitney U test (Hollander and Wolfe, 1973).

Results and discussion

Results of examinations

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):

not specified

Food efficiency:

not specified

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

not specified

Ophthalmological findings:

not specified

Haematological findings:

not specified

Clinical biochemistry findings:

not specified

Urinalysis findings:

not specified

Behaviour (functional findings):

not specified

Organ weight findings including organ / body weight ratios:

no effects observed

Gross pathological findings:

no effects observed

Histopathological findings: non-neoplastic:

no effects observed

Histopathological findings: neoplastic:

not specified

Details on results:

CLINICAL SIGNS AND MORTALITY
All mice survived to the end of the study. There were no chemical-related clinical findings.

BODY WEIGHT AND WEIGHT GAIN
The final mean body weights of exposed mice were similar to those of the control groups.

ORGAN WEIGHTS
There were no significant differences between control and exposed mice in absolute or relative organ weights.

GROSS PATHOLOGY
No chemical-related lesions were observed.

HISTOPATHOLOGY: NON-NEOPLASTIC
No chemical-related lesions were observed.

OTHER FINDINGS
There were no significant differences between control and exposed mice in epididymal weights, sperm count or motility.
There were significant increases in liver copper concentrations in female mice exposed to 30 mg/m^3 and in male and female mice exposed to 100 mg/m^3 compared to those of the control groups.

Effect levels

1

Dose descriptor:

NOAEC

Remarks:

for chemical related clinical findings or lesions

Effect level:

> 100 mg/m³ air (nominal)

Based on:

test mat.

Sex:

male/female

Basis for effect level:

other: see 'Remark'

2

Dose descriptor:

LOEC

Remarks:

for increased liver copper concentrations

Effect level:

30 mg/m³ air (nominal)

Based on:

test mat.

Sex:

female

Basis for effect level:

other: There were significant increases in liver copper concentrations in female mice exposed to 30 mg/m3 compared to those of the control groups.

3

Dose descriptor:

LOEC

Remarks:

for increased liver copper concentrations

Effect level:

100 mg/m³ air (nominal)

Based on:

test mat.

Sex:

male

Basis for effect level:

other: There were significant increases in liver copper concentrations in male mice exposed to 100 mg/m3 compared to those of the control groups.

Target system / organ toxicity

Critical effects observed:

not specified

Applicant's summary and conclusion

Conclusions:

No treatment-related effects on mortality, clincial signs, final mean body weights, organ weights, and epididymal weights, sperm count, or motility. No treatment-related gross or microscopic lesions were observed. Significant increases in liver copper concentrations were observed in females exposed to 30 mg/m^3 (by 15 %) and in males and females exposed to 100 mg/m^3 (by 40 % and 23 %, respectively) when compared to the controls.