Benzene-1,2:4,5-tetracarboxylic dianhydride

Repeated dose toxicity: inhalation

Administrative data

Endpoint:

short-term repeated dose toxicity: inhalation

Type of information:

experimental study

Adequacy of study:

key study

Study period:

03-03-1978 to 30-11-1978

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

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

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

1978

Report date:

1978

Materials and methods

Test guideline

Qualifier:

no guideline followed

Principles of method if other than guideline:

A scientifically acceptable short-term inhalation toxicity study in which rats were exposed to airborne dusts of the test material for 10 days. Appropriate study parameters were assessed and adequately reported.

GLP compliance:

not specified

Limit test:

no

Test material

Test material information

Constituent 1

Reference substance name:

Benzene-1,2:4,5-tetracarboxylic dianhydride

EC Number:

201-898-9

EC Name:

Benzene-1,2:4,5-tetracarboxylic dianhydride

Cas Number:

89-32-7

Molecular formula:

C10H2O6

IUPAC Name:

5,11-dioxatricyclo[7.3.0.0³,⁷]dodeca-1,3(7),8-triene-4,6,10,12-tetrone

Test material form:

solid: crystalline

Details on test material:

- Colour: colourless
- Odour: odourless
- CAS-Number: 89-32-7
- Molecular formula: C10 H2 O6
- Molecular weight: 218.12

Specific details on test material used for the study:

PMDA
pyromellitic dianhydride
IC-4054 3-72
Lot No. RI-18

Test animals

Species:

rat

Strain:

Sprague-Dawley

Details on species / strain selection:

Charles River CD rats, 6-8 weeks old

Sex:

male

Details on test animals or test system and environmental conditions:

Forty male (weighing from 196 to 228 grams) Charles River CD rats, 6-8 weeks of age were used in the study. The 40 males were randomized into 4 groups of 10 rats each. in accordance with a random table of 40. After the randomization process, the rats were individually identified with ear punches in numerical order from l to 40. Each rat was housed individually in compartmented stainless-steel holding cages and kept in a temperature and humidity-controlled room throughout the 1-week quarantine, the 2-veek experimental, and the 2-week post exposure observation periods. The rats were supplied with water and Purina rat Chow ad libitum except during exposure.

Administration / exposure

Route of administration:

inhalation: dust

Type of inhalation exposure:

whole body

Vehicle:

air

Mass median aerodynamic diameter (MMAD):

>= 3 - <= 3.7 µm

Remarks on MMAD:

GSD given as 2.32 to 2.43 in the study report.

Details on inhalation exposure:

All exposures were conducted in a 1 cubic meter cubical stainless steel and glass chamber, with pyramidal top and bottom. Chamber airflow was maintained by means of a rotary centrifugal air pump located at the exhaust side of the chamber. The chamber exhaust was filtered through an activated charcoal filter and a Cambridge Absolute filter before being discharged outside of the laboratory. The dust atmosphere of the compound was generated by dispersing the powder at a calculated rate with an IRAD Dust Gun located near the chamber air inlet at the top of the exposure chamber. This dust generator consisted of a revolving plate with calibrated cups for transporting a known quantity of powder per unit time from a. reservoir to a "blowhole.”. At the “blowhole" the powder in a "cup" was dispersed into the chamber by a jet of desiccated air flowing at a rate of 10 litres/minute.

Analytical verification of doses or concentrations:

yes

Details on analytical verification of doses or concentrations:

The actual quantity of powder disseminated was determined by weighing the quantity of powder in the reservoir before and after the exposure. The concentration of the dust in the chamber atmosphere was calculated from the ratio of the quantity of powder used to the total chamber airflow per unit time.
The concentration of the airborne dusts in the chamber atmosphere was determined gravimetrically using a fiberglass Millipore filter sampling technique. The chamber atmosphere was drawn through a 37 mm 0.8 µ pore size filter at a rate of 2 litres/minute. A critical orifice was used to regulate the flow race. Samples were taken from each exposure chamber once every 30 minutes during the exposure period. The weight of pyromellitic dianhydride collected on the filter was determined and the concentration of the duet was calculated in milligrams/litre of air.
The particle size distribution of the pyromellitic dianhydride in a sample of the chamber atmosphere was determined using an Andersen 8-stage Fractionating Sampler. The chamber atmosphere was drawn through the sampler at a rate of 28.3 1/min for a suitable duration. The weight of the particles collected on each stage is determined gravimetrically and the weight percent of each size category was calculated. The cumulative weight percent of particles smaller that each size range was plotted on a logarithmic probability graph paper. The aerodynamic mass median diameter and the percent of respirable particles in the sample was determined graphically by interpolation.

Duration of treatment / exposure:

Two weeks, 10 exposures, 6 hours per day.

Frequency of treatment:

10 exposures in 2 weeks.

Doses / concentrations

Doses / concentrations 1

Dose / conc.:

0 mg/L air

Remarks:

Control

Doses / concentrations 2

Dose / conc.:

0.01 mg/L air (nominal)

Remarks:

0.021 mg/L air analytical

Doses / concentrations 3

Dose / conc.:

0.03 mg/L air (nominal)

Remarks:

0.035 mg/L air analytical

Doses / concentrations 4

Dose / conc.:

0.1 mg/L air (nominal)

Remarks:

0.091 mg/L air analytical

No. of animals per sex per dose:

10 males

Control animals:

yes

Details on study design:

These are given in the previous sections.

Positive control:

No

Examinations

Observations and examinations performed and frequency:

The rats were observed daily before and after exposure for respiratory distress, nasal irritation and discharge, and any other observable toxic signs.

The rats were weighed daily on exposure days and periodically during the quarantine and post exposure observation period. Five rats from each group were sacrificed and necropsied following the 10th exposure. The remaining rats were observed for an additional 14 days and then sacrificed.

Sacrifice and pathology:

Five rats/group sacrificed after 10th exposure and the reamining rats after another 14 days.

Other examinations:

Clinical laboratory tests were conducted on 5 rats from each group at the termination of the exposure period, the remaining rats from each group were kept and observed for 14 days. At the end of the 14-day observation period, clinical laboratory tests were conducted on the remaining, 20 rats. For the collection of urine samples, the rats were placed in metabolism cages and fasted overnight. Blood was then collected by the orbital sinus puncture technique. The rats were then sacrificed for gross and histopathological examinations.
Haematological studies included erythrocyte count, total and differential leucocyte count, haemoglobin and haematocrit. Mean corpuscular volume and mean corpuscular haemoglobin were calculated.
Biochemical studies included serum glutamyl transpeptidase, alkaline phosphatase, bilirubin, lactic dehydrogenase, serum glutamic oxaloacetate and pyruvic transaminase activities, blood urea nitrogen and total protein.
Urinalyses included measurement of volume, osmolality, pH, sugar, proteins , blood, urobilinogen, bilirubin and microscopic examination of sediment.

Statistics:

All statistical analyses compared the treatment groups with the control group.
Erythrocytes, haemoglobin, mean corpuscular volume, mean corpuscular haemoglobin, MCRC, LDH (post 10 exposure and 13 day post exposure) aid absolute and relative organ weights withdrawal sacrifice and terminal sacrifice) were compared by analysis of variance (one-way classification), Bartlett's test for homogeneity of variances and the appropriate t-test (for equal or unequal variances) using Dunnett’s multiple comparison tables to judge significance of differences.

Results and discussion

Results of examinations

Clinical signs:

effects observed, treatment-related

Description (incidence and severity):

The most notable observable changes in the rats after exposure to the compound were respiratory difficulties and the appearance of nasal irritation and discharge. There was an observable gradient in the severity and onset of these effects from the low to high exposure levels. By the end of the third day the rats exposed to 0.01 mg/1experienced slight breathing difficulties and nasal irritation. The rats exposed to 0.03 mg/1 of the dust exhibited nasal irritation with a few intermittent cases of red nasal discharge. By the end of the third exposure day the rats experienced markedly laboured respiration. The rats in the 0.1 mg/1 exposure group had nasal and respiratory involvement from the first day of exposure. Nasal irritation had progressed to a red nasal discharge in 50-80% of these rats. By the second exposure day all rats in this group had marked difficulty in breathing, about half to the point of gasping.

Mortality:

no mortality observed

Body weight and weight changes:

effects observed, treatment-related

Description (incidence and severity):

The body weights recorded before, during and after the exposure period showed a direct relationship between body weight loss and exposure concentrations. The control rats, which were exposed to filtered air, gained weight throughout the experimental period. The rats at 0.01 and 0.03 mg/1 maintained weight during exposure and gained weight on weekends when they were not exposed. The rats exposed to 0.1 mg/1 lost weight during the exposure periods but showed weight gains on the weekends. At the termination of the 10 exposures, the mean body weights of the rats of the 0.01 and the 0.03 mg/1 groups were below that of the control group but were above the pre-exposure mean weights. The mean body weight of the rats in the 0.1 mg/l group at the end of the exposure period was below the pre-exposure mean weight.

All the exposed rats, which were kept for post exposure observations, showed body weight gain. At the end of the observation period, the mean body weights of all the exposed rats were still below that of the control group in a concentration related manner.

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):

Examination of the hematological parameters of rats which were sacrificed shortly after the 10th exposure revealed a significant (p<0.05) increase in the number of erythrocytes per unit volume of blood in the 0.03 and 0.1 mg/l concentration groups and a significant (p<0.05 and 0.01) increase in haemoglobin concentration in all the exposed groups. A significant (p<0.01) increase in neutrophils also was seen for rats at the 0.1 mg/1 dosage level. Similar examinations of the rats which were sacrificed 2 weeks after the exposures revealed no significant differences between the groups.

Clinical biochemistry findings:

effects observed, treatment-related

Description (incidence and severity):

Examination of the data revealed a significant (p<0.01) depression of the level of lactate dehydrogenase in the rats which were sacrificed immediately after the exposure. to 0.1 mg/1 of the compound. However, similar examinations of the rats, which were sacrificed after 14 days of observation revealed no depression of LDH in all groups. There were no other detectable changes seen in the studies of serum alkaline phosphatase, glutamic pyruvic transaminase and gamma glutamyl transpeptidase activities and the levels of blood urea nitrogen and total protein at all time intervals.

Urinalysis findings:

no effects observed

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):

Some variations were noted but were of unknown biologic significance.
There was an increase in lung weight noted at the 0.01 mg/1 - 0.1 mg/l level, but this was unaccompanied by morphologic alterations.

Gross pathological findings:

no effects observed

Description (incidence and severity):

No gross lesions considered compound-related were seen in rats sacrificed at tile end of exposure and those sacrificed after a compound withdrawal period of 14 days.

Neuropathological findings:

not examined

Histopathological findings: non-neoplastic:

effects observed, treatment-related

Description (incidence and severity):

No microscopic lesions considered compound-related were observed in tissues other than lungs of rats from the test groups at either sacrifice.

All rats in the 0.01, 0.03 and 0.1 mg/1 exposure groups sacrificed at the end of the exposure period or after 14 days of compound withdrawal had very slight to slight multifocal hypertrophy and hyperplasia of type 2 alveolar cells with occasional formation of alveolar giant cells (foreign body type). Most rats had very slight to slight focal to multifocal interstitial pneumonia characterized by very slight to slight interstitial fibrosis and very slight to slight interstitial mononuclear inflammatory cell infiltrates. All rats in the 3 test groups sacrificed after 10 exposures had very slight to slight aggregates of foamy macrophages in bronchiolar lumens; two rats, one from the 0.01 mg/1 group and one from the 0.03 mg/1 group, at 14th day post exposure had very slight aggregates of foamy macrophages in bronchiolar lumens. All rats in the 3 test groups at 14th day post exposure had slight to very slight aggregates of dark-brown pigment in cytoplasm of hypertrophic and hyperplastic type 2 alveolar cells; no similar pigment was observed in rats from these groups sacrificed immediately after 10 exposures. No microscopic changes which were considered treatment related were seen in the lungs of rats from the control group of either sacrifice.

Histopathological findings: neoplastic:

no effects observed

Other effects:

no effects observed

Effect levels

Key result

Dose descriptor:

LOAEL

Remarks:

local effects

Effect level:

>= 0.021 - <= 0.091 mg/L air

Based on:

test mat.

Sex:

male

Basis for effect level:

clinical signs

organ weights and organ / body weight ratios

Remarks on result:

other: Slight/very slight changes seen only in the lungs

Target system / organ toxicity

Critical effects observed:

no

Applicant's summary and conclusion

Conclusions:

Rats exposed to dust atmospheres of pyromellitic dianhydride for 6 hours per day for a total of 10 exposures in 2 weeks (nominal: 0.01. 0.03 and 0.1mg/L; analysed: 0.021, 0.035 and 0.091 mg/L) exhibited symptoms directly related to the route of exposure, i.e. most effects were evident in the lungs. The signs were indicative of local, direct effects in the lungs. PMDA is highly reactive and hydrolyses in contact with water or body fluids to the corresponding pyromellitic acid; PMDA is also a severe eye irritant. It is unsurprising that direct local effects in the lungs were observed. Other changes seen in this study were likely to be related to physiological stress because of the clinical signs induced via inhalation exposure to the dusts. Only the lungs showed any histopathological treatment related signs, importantly the signs seen were only slight to very slight.

Executive summary:

Male rats were exposed to dust atmospheres of pyromellitic dianhydride for 6 hours per day for a total of 10 exposures in 2 weeks. Ten rats were used at nominal dosage levels of 0.01. 0.03 and 0.1mg/l and in a control group. Rats exposed to a concentration of 0.1 mg/1 exhibited a red nasal discharge and breathing difficulties to the point of gasping. The group exposed to 0.03 mg/1 had laboured breathing and nasal irritation with a few incidences of red nasal discharge. At an exposure level of 0.01 mg/1 the rats had slight breathing difficulties and nasal irritation. The severity of the laboured breathing and nasal effects decreased with decreasing concentration. All exposed groups had decreased weight gains. Examination of the hematological parameters of rats which were sacrificed shortly after the 10th exposure revealed an increase in the number of erythrocytes per unit volume of blood in the 0.03 and 0.1 mg/1 concentration groups and an increase in haemoglobin concentration in all the exposed groups. An increase in neutrophils also was seen for rats at the 0.1 mg/1 dosage level. Lactate dehydrogenase activity was reduced in the rats which were sacrificed immediately after the exposures to 0.1 mg/1 of the compound. No gross lesions or organ weight variations considered compound related were seen in rats sacrificed after 10 exposures and those sacrificed after a compound withdrawal period of 14 days. No microscopic pathological lesions which were considered compound related were seen in tissues other than the lungs of rats at the 0.01, 0.03 and 0.1 mg/1 exposure levels sacrificed after 10 exposures and after 14 days withdrawal. microscopically, the lung lesions considered compound related were very slight to slight, focal to multifocal interstitial pneumonia, very slight to slight aggregates of foamy macrophages in bronchiolar lumens, and very slight to slight multifocal hypertrophy and hyperplasia of type 2 alveolar cell, with occasional formation of alveolar giant cells (foreign body type).

In conclusion, rats exposed to dust atmospheres of pyromellitic dianhydride for 6 hours per day for a total of 10 exposures in 2 weeks (nominal: 0.01. 0.03 and 0.1mg/L; analysed: 0.021, 0.035 and 0.091 mg/L) exhibited symptoms directly related to the route of exposure, i.e. most effects were evident in the lungs.  The signs were indicative of local, direct effects in the lungs. PMDA is highly reactive and hydrolyses in contact with water or body fluids to the corresponding pyromellitic acid; PMDA is also a severe eye irritant. It is unsurprising that direct local effects in the lungs were observed. Other changes seen in this study were likely to be related to physiological stress because of the clinical signs induced via inhalation exposure to the dusts. Only the lungs showed any histopathological treatment related signs, importantly the signs seen were only slight to very slight.