Indium phosphide

Repeated dose toxicity: inhalation

Administrative data

Endpoint:

sub-chronic toxicity: inhalation

Type of information:

experimental study

Adequacy of study:

key study

Study period:

3-4 April 1995 to 4-5 July 1995

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

comparable to guideline study with acceptable restrictions

Data source

Reference

Reference Type:

publication

Title:

Unnamed

Year:

2001

Materials and methods

Test guideline

Qualifier:

equivalent or similar to guideline

Guideline:

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

Deviations:

yes

Remarks:

Several organs not weighed,sperm motility and vaginal cytology not performed on animals exposed to the highest concentration,some examinations notably food consumption not included

GLP compliance:

yes

Limit test:

no

Test material

Test material information

Constituent 1

Reference substance name:

Indium phosphide

EC Number:

244-959-5

EC Name:

Indium phosphide

Cas Number:

22398-80-7

Molecular formula:

InP

IUPAC Name:

phosphinidyneindium

impurity 1

Reference substance name:

Impurities of BNW 12957-28 [14 week]

Molecular formula:

N/A

IUPAC Name:

Impurities of BNW 12957-28 [14 week]

impurity 2

Reference substance name:

Other Impurities

Molecular formula:

N/A

IUPAC Name:

Other Impurities

Test material form:

solid: crystalline

Test animals

Species:

rat

Strain:

Fischer 344

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source:Taconic Farms (germantown,NY)
- Females nulliparous and non-pregnant: Yes
- Age at study initiation: 6 weeks
- Weight at study initiation:male: average = 124.6g (average) female: average= 106.3g (average)
- Housing: Individually in stainless steel wire bottom cages.
- Diet: NTP-2000 open formula pelleted diet ad libitum (except during exposure periods)
- Water: Softened tap water ad libitum (except during exposure and urine collection periods)
- Acclimation period: 12 days males or 13 days females

ENVIRONMENTAL CONDITIONS
- Temperature (°C):24° +/- 3
- Humidity (%):55% +/-15%
- Air changes (per hr):Chamber air changes 15/hr
- Photoperiod (hrs light): 12 hours per day

Administration / exposure

Route of administration:

inhalation: aerosol

Type of inhalation exposure:

whole body

Vehicle:

air

Mass median aerodynamic diameter (MMAD):

>= 1.2 - <= 1.4 µm

Geometric standard deviation (GSD):

1.6

Details on inhalation exposure:

GENERATION OF TEST ATMOSPHERE / CHAMBER DESCRIPTION
- Exposure apparatus: Stainless-steel inhalation exposure chambers (Harford Systems Division of Lab Products, Inc., Aberdeen, MD)
- Source and rate of air: Compressed air
- Method of conditioning air: The aerosol leaving the generator passed through a corona discharge air-ionizing neutralizer into the distribution line. At each chamber location, a pneumatic injector drew aerosol from the distribution line into the chamber inlet, where the aerosol was further diluted with HEPA-filtered air to the appropriate concentration.
- System of generating particulates/aerosols: the aerosol generation and delivery system consisted of a flexible brush dust feed mechanism, a Trost model GEM-T air impact mill,an aerosol charge neutralizer and a stainless steel aerosol distribution system.The flexible-brush dust feed mechanism contained a hopper into which dry powder was poured. The hopper was reloaded at regular intervals throughout each day’s exposure period. Indium Phosphide was stored in a nitrogen purged desiccator to gain a more uniformed flow in the generator.
- Temperature, humidity, pressure in air chamber: Temperature (°C): 24° +/- 3 Humidity (%):55% +/-15%
- Air change rate: 15 air changes per hour
- Method of particle size determination: Mercer-style seven-stage impactor.Stages analyzed by inductively coupled plasma/mass spectroscopy (ICP/AES). Relative mass collected on each stage was analyzed by probit analysis. The mass median aerodynamic particle diameter and the geometric standard deviation of each set of samples were estimated.

TEST ATMOSPHERE
- Brief description of analytical method used: Mercer-style seven-stage impactor (In-Tox Products, Albuquerque, NM). The stages (glass coverslips lightly sprayed with silicone) were analyzed by inductively coupled plasma/mass spectroscopy (ICP/AES).
- Samples taken from breathing zone: yes

Analytical verification of doses or concentrations:

yes

Details on analytical verification of doses or concentrations:

Chamber aerosol concentrations were monitored with real-time aerosol monitors using a pulsed-light-emitting diode in combination with a silicon detector to sense light scattered over a forward angular range of 45° to 95° by particles traversing the sensing volume.

Duration of treatment / exposure:

14 weeks

Frequency of treatment:

0, 1, 3, 10, 30, or 100 mg/m3, 6 hours plus T90 (12 minutes) per day, 5 days per week (weeks 1 to 4 and weeks 10 to 14) or 7 days per week (weeks 5 to 9) during the concurrent teratology study

Doses / concentrations

Doses / concentrations 1

Dose / conc.:

0 mg/m³ air

Doses / concentrations 2

Dose / conc.:

1 mg/m³ air

Doses / concentrations 3

Dose / conc.:

3 mg/m³ air

Doses / concentrations 4

Dose / conc.:

10 mg/m³ air

Doses / concentrations 5

Dose / conc.:

30 mg/m³ air

Doses / concentrations 6

Dose / conc.:

100 mg/m³ air

No. of animals per sex per dose:

10 animals/sex/dose

Control animals:

yes, concurrent no treatment

Examinations

Observations and examinations performed and frequency:

CAGE SIDE OBSERVATIONS: Yes
- Time schedule: Twice daily

DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: week 1, weekly, and at the end of the study.

BODY WEIGHT: Yes
- Time schedule for examinations: Initially, weekly, and at the end of the study.

FOOD CONSUMPTION AND COMPOUND INTAKE (if feeding study): No

FOOD EFFICIENCY: No

WATER CONSUMPTION AND COMPOUND INTAKE (if drinking water study): No

OPHTHALMOSCOPIC EXAMINATION: No

HAEMATOLOGY: Yes
- Time schedule for collection of blood: day 3, 23, week 14
- Anaesthetic used for blood collection: Yes 70% CO2/air mixture
- How many animals: 10 animals per sex/ per dose (with exception) Week 14 100mg/m3 male group used 9 rats.
- Parameters checked in table No.2 were examined.

CLINICAL CHEMISTRY: Yes
- How many animals: 10 animals per sex/ per dose (with exception) Week 14 100mg/m3 male group used 9 rats.
- Parameters checked in table No.2 were examined.

URINALYSIS: Yes
- Time schedule for collection of urine: day 31
- Metabolism cages used for collection of urine: Yes
- Animals fasted: No water during urine collection period
- Parameters checked in table No.2 were examined.

NEUROBEHAVIOURAL EXAMINATION: No

IMMUNOLOGY: No

OTHER:
Necropsy: Yes
- Time schedule for examinations: Study Termination
- How many animals: all core study animals
- Dose groups that were examined:all core study animals
- Parameters checked in table No.2 were examined.

Sperm Motility and Vaginal Cytology: Yes
-Time schedule for examinations: Study termination
-Dose groups that were examined: Sperm samples were collected from male rats 0, 3, 10, and 30 mg/m3 groups
- Parameters checked in table No.2 were examined.

Tissue Burden Studies: Yes
- Time schedule for examinations: day 4, 24, 45, 73, or 96
- How many animals: three
- Dose groups that were examined: per group
- Parameters checked in table No.2 were examined.

Postexposure Lung Burden Study: Yes
- Time schedule for examinations: Day 14, 28, 56, or 112.
- How many animals: three to five rats
- Dose groups that were examined:per group
- Parameters checked in table No.2 were examined.

Postexposure Lung Burden Study in Age-Matched Animals: Yes
- Time schedule for examinations: 5,14, 28, 56, or 112 days
- How many animals: 3 rats
- Dose groups that were examined: per group
- Parameters checked in table No.2 were examined.

Sacrifice and pathology:

GROSS PATHOLOGY: Yes see table No. 2
HISTOPATHOLOGY: Yes see table No. 2

Statistics:

Organ and body weight data were analysed with the parametric multiple comparison procedures of Dunnett (1955) and Williams (1971, 1972). Haematology, clinical chemistry, urinalysis, spermatid, and epididymal spermatozoal data, were analysed using the nonparametric multiple comparison methods of Shirley (1977) and Dunn (1964). For neoplasms and non-neoplastic lesions detected, the Fisher exact test (Gart et al., 1979) was used.

Results and discussion

Results of examinations

Clinical signs:

effects observed, treatment-related

Description (incidence and severity):

Shallow, rapid, abnormal breathing observed in males and females exposed to 30 or 100mg/m3. Animals in the 100mg/m3 groups exhibited letharygy, thinness and ruffled fur.

Mortality:

mortality observed, non-treatment-related

Description (incidence):

One male in the 100mg/m3 died before the end of the study.

Body weight and weight changes:

effects observed, treatment-related

Description (incidence and severity):

Mean body weights and body weight gains of males and females in the 100mg/m3 exposed groups were significantly less than those of the chamber controls.

Food consumption and compound intake (if feeding study):

not examined

Food efficiency:

not examined

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

not examined

Ophthalmological findings:

not examined

Haematological findings:

effects observed, treatment-related

Description (incidence and severity):

On day 23 and week 14 all groups of exposed male and female rats showed signs of exposure concentration-related erythrocytosis. At week 14 erythrocytosis was accompanied by increased reticulocyte and nucleated erythrocyte cell counts in the 100mg/m3 groups. At week 14 mean cell volume, mean cell haemoglobin and mean cell haemoglobin concentration showed concentration related decreases in the 10mg/m3 or greater female and or male groups signifying that for the high-exposure animals, the circulating erythrocytes were smaller and contained less hemoglobin than expected. On days 3 and 23 there was evidence of exposure related decrease in leukocyte counts. By day 23 this effect was seen in all exposed groups of male and in the 30 and 100mg/m3 groups of females. At week 14 leukocyte counts increased in the 10mg/m3 males and the 30 and 100 mg/m3 females. Exposure related increases of leukocyte counts were ascribed to pulmonary inflammation. Platelet count decreases were noted in numerous higher exposure groups however they were not considered to be clinically or toxicologically relevant.

Clinical biochemistry findings:

effects observed, treatment-related

Description (incidence and severity):

On day 23 and week 14 increases in serum alanine aminotransferase and sorbitol dehydrogenase activities showed a hepatocellular effect. By week 14 increased alanine aminotransferase and sorbitol dehydrogenase activities occured in 10mg/m3 or greater females and in all groups of exposed males.

Urinalysis findings:

effects observed, treatment-related

Description (incidence and severity):

At week 14, decreased total protein, albumin and creatinine and increased urea nitrogen concentrations occured in all the 100mg/m3 groups of males and females which was considered to be consistent with decreased weight gain in these groups and was though to be due to a compromised nutritional status.

Behaviour (functional findings):

not examined

Immunological findings:

not examined

Organ weight findings including organ / body weight ratios:

effects observed, treatment-related

Description (incidence and severity):

Lung weights of all exposed groups of both male and female rats were proportionate to the increase in exposure and were significantly higher than those of the chamber controls. A 2.7 to 4.4- fold increase in the absolute lung weights was affiliated to the build up of proteinaceous fluid (alveolar proteinosis) in the alveoli. Relative heart weights were also increased in the 30 and 100mg/m3 exposed groups of males and females, and absolute heart weights were increased in the 10 mg/m3 or greater females. Thymus weights were decreased in the males and females exposed to 100mg/m3 when compared to chamber controls, these decreases were thought to be related to the weakened state of the rats.

Gross pathological findings:

effects observed, treatment-related

Description (incidence and severity):

Gross exposure related lesions were observed in the lungs which increased in severity with increasing exposure concentration. Lungs of all exposed rats were enlarged and had a gray to black discoloration and a granular to dimpled appearance.Chronic active inflammation of the lung occurred in all exposed animals and was multifocal to diffuse. Inflammation was composed of mixed inflammatory cells within the alveoli and interstitium. Within areas of inflammation, there was regenerative alveolar epithelial hyperplasia whose severity increased with increasing exposure concentration in males. In the nose, larynx, and trachea of exposed males and females, minimal to mild build-up of foreign bodies (indium phosphide particles) occurred in the mucosal epithelial cells or the underlying substantia propria, either free or within macrophages. Furthermore, in the base of the epiglottis of male and female rats in the 3, 10, or 30 mg/m3 groups, there were marginally severe groups of mononuclear cells (inflammation) associated with the particles.

Neuropathological findings:

not examined

Histopathological findings: non-neoplastic:

effects observed, treatment-related

Description (incidence and severity):

Hyperplasia is typical in local lymph nodes draining areas of foreign material deposition and inflammation. The Hyperplasia recorded was multifocal and was composed of differentiated cuboidal epithelial cells. Interstitial fibrosis occurred in most males and females exposed to 3mg/m3 or greater and severities were proportionate to increasing concentrations. The majority of bronchial and mediastinal lymph nodes observed from exposed males and females were enlarged and contained augmented numbers of lymphocytes and larger immature mononuclear cells. Marginal differences in the severities of hyperplasia among exposed groups were observed. Pigmentation (indium phosphide particles) also occurred in the lymph node macrophages of most exposed animals, the quantities increased with increasing exposure concentration. Incidences of moderately severe hyperplasia of the bone marrow and mild hematopoietic cell proliferation of the spleen were increased in males and females exposed to 100 mg/m3. The incidence of renal nephropathy was significantly increased in 100 mg/m3 females. Minimal nephropathy occurred in chamber control in 1, 3, 10, and 30 mg/m3 rats, but the severity was moderate in 100 mg/m3 males and mild in 100 mg/m3 females. Progressive degenerative nephropathy occurs spontaneously in F344/N rats, is generally more severe in males, and is often aggravated by chemical exposure. Centrilobular atrophy and centrilobular necrosis of the liver were significantly increased in male and female rats exposed to 100 mg/m3. Hemosiderin pigmentation was increased in females exposed to 100 mg/m3. It could not be determined if the centrilobular atrophy and necrosis were primary toxic effects or were secondary to hypoxia resulting from the severe lung lesions. The incidence of hepatodiaphragmatic nodules was marginally increased in female rats exposed to 100 mg/m3, the reason for the increased incidence was not determined and the occurrence was not considered biologically significant. Increased incidences of hypertrophy of the heart in male and female rats exposed to 100 mg/m3 were recorded. The incidences of a numerous lesions were significantly increased only in the 100 mg/m3 rats and were considered secondary to debilitation. No significant differences were noted in sperm morphology or vaginal cytology parameters between exposed and chamber control rats that could be attributed to a direct effect of indium phosphide exposure.

Histopathological findings: neoplastic:

not examined

Other effects:

effects observed, treatment-related

Description (incidence and severity):

Tissue burden analyses were performed on male rats in the 14-week study and age-matched male rats exposed with the 14-week study animals for the last five days of the 14-week study. Compared to chamber control animals, lung weights of exposed male rats increased throughout the 14-week exposure period and continued to increase throughout the 16-week recovery period with the exception of animals in the 1mg/m3 groups whose lung weights remained relatively unchanged through the 16-week recovery period. Lung weights of rats exposed to 3, 10, or 30 mg/m3 were comparable during the exposure and recovery periods. Although lung weights of rats exposed to 100 mg/m3 were greater than those of other exposed groups on day 4, they were similar to the lower exposure concentration groups during the study, and were lower than the other exposed groups towards the end of the 14 weeks of exposure. Lung weights of age-matched male rats exposed for 5 days increased with increasing exposure concentration and continued to increase significantly more than the chamber control animal lung weights throughout the 16-week recovery period. At the end of the recovery period, lung weights were significantly increased; however, the lung weights of the age matched rats were considerably less than those of rats exposed continuously for 14 weeks.

Lung burdens of indium increased with increasing exposure concentration and each increased throughout the 14 weeks of exposure demonstrating that steady-state lung burdens for indium were not attained. Lung burdens for the 14 weeks of exposure and the succeeding 16-week recovery period were standardised to exposure concentration to measure their proportionality to exposure concentration. Data showed that through the 14-week exposure period and the subsequent recovery period, lung burdens were excessively low for the 30 and 100 mg/m3 groups when compared to the 10 mg/m3 or lower groups. Calculated lung clearance half-times during the 14-week exposure period were not significantly different between exposed groups. Although lung deposition rates increased with increasing exposure concentration, lung deposition rates normalized to exposure concentrations decreased with increasing exposure concentration. Therefore, at the higher exposure concentration, the amount of indium deposited per unit exposure concentration was less than at lower concentrations. This was thought to be due to an alteration in the pulmonary function caused by inflammatory and proliferative lesions in the lung. Lung burdens also decreased for postexposure animals with time. The calculated lung clearance rate constants or halftimes determined during the 16-week recovery period were not different for the 1, 3, 10, or 30 mg/m3 groups. The normalized lung burdens of age-matched rats in the 5-day study showed that the lung burdens were proportionate to exposure concentration excluding the 30 and 100 mg/m3 groups at the initial time point and for the 100 mg/m3 group at 16 weeks postexposure. Normalized deposition rates calculated for the 5-day exposure were not different between exposure groups or from those calculated from lung burdens determined at 4 days of exposure during the 14-week study. Therefore, lung deposition did not vary as a consequence of age. However, the normalized deposition rates from the 5-day exposure were approximately two-fold higher than those calculated from the 14-week study data, signifying that prolonged exposure to indium phosphide may have caused decreases in deposition rates. The overall mean lung clearance halftimes after the 5-day exposure averaged 146+/- 68 days, halfway amid the clearance half-times measured during the 14-week exposure and 16-week postexposure periods, and were not different from either.

Effect levels

Key result

Dose descriptor:

LOAEC

Effect level:

1 mg/m³ air

Based on:

test mat.

Sex:

male/female

Basis for effect level:

gross pathology

haematology

histopathology: non-neoplastic

immunology

organ weights and organ / body weight ratios

Target system / organ toxicity

1

Key result

Critical effects observed:

yes

Lowest effective dose / conc.:

1 mg/m³ air

System:

respiratory system: lower respiratory tract

Organ:

alveoli

lungs

lymph node

Treatment related:

yes

Dose response relationship:

yes

Relevant for humans:

not specified

2

Key result

Critical effects observed:

no

Lowest effective dose / conc.:

1 mg/m³ air

System:

respiratory system: upper respiratory tract

Organ:

larynx

nasal cavity

trachea

Applicant's summary and conclusion

Conclusions:

Haematological, immunological, gross pathological and lower respiratory tract adverse effects were identified during a subchronic toxicity study conducted on rats via the inhalation route.

Executive summary:

A GLP-compliant study using a method similar to the OECD TG 413 (with deviations) was conducted to evaluate the repeated dose toxicity potential of indium phosphide via inhalation.

Groups of 10 Male and 10 female F344/n rats were exposed to particulate aerosols of indium phosphide at concentrations of 0, 1, 3, 10, 30, or 100 mg/m3 for 6 hours per day, 5 days per week (weeks 1 to 4 and weeks 10 to 14) or 7 days per week (weeks 5 to 9).

Examinations included daily observations (twice), weekly body weight and clinical sign recordings, mortality, body weights changes, haematology, clinical chemistry, organ weigh findings, urinalysis, histopathology, sperm motility, vaginal cytology, and burden study on lung, testis, blood and serum from male rats were recorded. One male in the 100 mg/m3 group died before the end of the study.

At the end of the study, surviving animals were subjected to gross necropsy and histopathology. Mean body weights and body weight gains of all exposed groups in the 100 mg/m3 group were significantly less than the chamber controls. The 100 mg/m3 exposed groups exhibited lethargy, thinness and ruffled fur as well as abnormal breathing in the 30 mg/m3 groups. All exposed male and female rats showed signs of exposure concentration-related erythrocytosis. In the 100 mg/m3 groups increased reticulocyte and nucleated erythrocyte cell counts were recorded. In the 10 mg/m3 groups concentration related decreases in erythrocyte indices were present. A decrease in leukocyte counts was seen in all exposed groups of male rats and in the 30 and 100 mg/m3 groups of female rats at day 3 and 23. At week 14 leukocyte counts increased in the 10 mg/m3 males and the 30 and 100 mg/m3 females. Increased alanine aminotransferase and sorbitol dehydrogenase activities occurred in 10 mg/m3 or greater females and in all groups of exposed males. Chronic active inflammation of the lung occurred in all exposed animals. Build-up of indium phosphide particles occurred in the nose, larynx, and trachea of exposed males and females. Lung weights of both male and female rats were proportionate to the increase in exposure and were significantly higher (A 2.7 to 4.4- fold increase) than those of the chamber controls. Relative heart weights increased in the 30 and 100 mg/m3 exposed groups of, where absolute heart weights increased in the >10 mg/m3 females. Thymus weights were decreased in the males and females exposed to 100 mg/m3 when compared to chamber controls, these decreases were thought to be related to the weakened state of the animals. Interstitial fibrosis occurred in most males and females exposed to 3 mg/m3 or greater and severities were proportionate to increasing concentrations. The majority of bronchial and mediastinal lymph nodes observed from exposed males and females were engorged containing amplified numbers of lymphocytes and larger immature mononuclear cells. Incidences of moderately severe hyperplasia of the bone marrow and mild hematopoietic cell proliferation of the spleen were increased in males and females exposed to 100 mg/m3. The incidence of renal nephropathy was significantly increased in 100 mg/m3 females. Minimal nephropathy occurred in chamber control in 1, 3, 10, and 30 mg/m3 rats, but the severity was moderate in 100 mg/m3 males and mild in 100 mg/m3 females. No significant differences were noted in sperm morphology or vaginal cytology parameters between exposed and chamber control rats that could be attributed to a direct effect of indium phosphide exposure. Tissue burden analyses were performed on male rats in the 14-week study and age-matched male rats exposed with the 14-week study animals for the last five days of the 14-week study. At the end of the recovery period, lung weights were significantly increased. Lung burdens of indium increased with increasing exposure concentration and each increased throughout the 14 weeks of exposure demonstrating that steady-state lung burdens for indium were not attained. While lung deposition rates increased with increasing exposure concentration, lung deposition rates normalized to exposure concentrations decreased with increasing exposure concentration. Therefore, at higher exposure concentrations the amount of indium deposited per unit exposure concentration was less than at lower concentrations. Overall mean clearance half-time was longer when calculated using the post exposure data (202 +/- 44 days) than when calculated from the 14- week exposure data (78 +/- 24 days). The overall mean lung clearance half-times after the 5 -day exposure averaged 146 +/- 68 days. Indium was detected in blood and serum at concen­trations less than that observed in lung tissue. Indium in the testis was detected at much higher concentrations than in blood or serum but less than that in the lung. Correspondingly, testicular indium concentration increased with increasing exposure concentration throughout the exposure period. Unlike blood and serum indium concentrations, testicular indium continued to increase in all groups following exposure, indicating that indium was accumulating in the testis over time.