Captan

Repeated dose toxicity: inhalation

Administrative data

Endpoint:

sub-chronic toxicity: inhalation

Type of information:

experimental study

Adequacy of study:

key study

Study period:

17 October 1988 to 19 January 1989

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

1989

Report date:

1989

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

EU Method B.29 (Sub-Chronic Inhalation Toxicity:90-Day Study)

Deviations:

yes

Remarks:

Temperature and humidity levels were occasionally outside nominal range. One male and one female (both 12.98 μg/L) were returned to the wrong holding cage, resulting in pregnancy. The pregnant animal was subsequently terminated.

Test guideline 2

Qualifier:

according to guideline

Guideline:

EPA OPP 82-4 (90-Day Inhalation Toxicity)

Deviations:

yes

Remarks:

Temperature and humidity levels were occasionally outside nominal range. One male and one female (both 12.98 μg/L) were returned to the wrong holding cage, resulting in pregnancy. The pregnant animal was subsequently terminated.

GLP compliance:

yes

Limit test:

yes

Test material

Test material information

Constituent 1

Reference substance name:

Captan

EC Number:

205-087-0

EC Name:

Captan

Cas Number:

133-06-2

Molecular formula:

C9H8Cl3NO2S

IUPAC Name:

2-[(trichloromethyl)sulfanyl]-2,3,3a,4,7,7a-hexahydro-1H-isoindole-1,3-dione

Test material form:

solid: particulate/powder

Details on test material:

- Name of test material: Captan, N-(trichloromethylthio)cyclohex-4-ene-1,2-dicarboximide
- Molecular formula: C9H8Cl3NO2S
- Molecular mass: 300.59
- Physical state: white powder
- Analytical purity: 88.7% (w/w)
- Batch number: 11240-37-1

Test animals

Species:

rat

Strain:

Wistar

Sex:

male/female

Details on test animals or test system and environmental conditions:

- groups of 40 male and 40 female specific pathogen free, A1pk:APfSD (Wistar derived), albino rats
- Source: Barriered Animal Breeding Unit (BABU) Alderley Park. Cheshire. UK
- age: five to six weeks
- free access to mains drinking water and food, except during exposure periods when all food and water was removed.
- room temperature: 20-24°C
- relative humidity: 40-60%

Administration / exposure

Route of administration:

inhalation: dust

Type of inhalation exposure:

nose only

Vehicle:

air

Details on inhalation exposure:

Animals were exposed nose-only in restraining tubes supplied by Battelle, Geneva. Switzerland, which were Inserted into an ICl designed PERSPEX exposure chamber having an internal volume of approximately 46 litres. Animals were placed in the exposure chambers in replicate order with male and females on separate levels. Their exact position on each level was varied dally as was the level on which they were exposed. The exposure period commenced once the restraining tubes were inserted into the chambers and atmosphere generation had started. After six hours atmosphere generation was stopped and the animals were removed starting from the lowest group number. Temperature and relative humidity within each chamber during exposure were measured at approximately 30 minute intervals using a Vaisala HMI 31 portable digital temperature and humidity monitor (supplied by Vaisala UK Ltd. Cambridge, UK). Temperature and relative humidity were generally within the range of 22 ±3°C and 50 ±15% respectively. Some variation was seen in the latter, mainly during the earlier stages of the study and more frequently at the highest concentration but these are not considered to have affected the outcome of the study.Trial generations were carried out during a preliminary study. Atmospheres were generated into a reservoir chamber using a Wright's dust feed mechanism. A glass concentric jet atomiser was used above each exposure to create a venturi, which pulled test atmosphere from the reservoir chamber into the exposure chamber. Clean, dry air (dried and filtered using equipment supplied by Atlas-Copco, Sweden) was supplied to the exposure chamber via the atomiser and also directly, as diluting air. Airflow rates were measured using variable area flowmeters (KDG Flowmeters, Burgess Hill, Sussex, UK}. The airflow rates supplied to the atomisers were nominally: Group 2: 4.3 L/min (range 2.8-5.0 L/min), Group 3: 3.0 L/min (range 2.6-4.3 L/min), Group 4: 5.0 L/min (range 4.0-6.0 L/mln), Group 5: 16.0 L/mln (range 14.0-17.0 L/min}, Dilution air was supplied direct to the exposure chambers at the following flow rates (corrected to normal temperature and pressure): Group 1: 30 L/mln (range 29-30 L/mln), Group 2: 25 L/min (range 23-29 L/mln), Group 3: 25 L/mln (range 21-26 L/mln) Days 3 to 42: Group 4: 15 L/mln (range 15-15 L/min). Days 45 to 95: Group 4: 20 L/mln (range 19-25 L/mln). Days 3 to 42: Group 5: 0 L/mln. Days 45 to 95: Group 5: 8 L/mln (range 6-8 L/mln). At day 45 the volumes of diluting air supplied to the chambers of group 4 and 5 were altered. This was carried out in order to Increase the relative humidity of the air supplied to these chambers. This demonstrated that there were no significant differences in concentration between the top and bottom, and the front and back of the chamber.

Analytical verification of doses or concentrations:

yes

Details on analytical verification of doses or concentrations:

Atmospheres were sampled close to the animals breathing zone for determination of captan. The test atmosphere was drawn at a flow rate of 2 L/min over a measured time period, through a 25 mm diameter Vinyl Metrlcel (VN-I) filter housed in a Delrin open-faced filter holder (filters and holders supplied by Gelman Sciences Ltd., Northampton, UK).
The filter was weighed before and after sampling and the concentration was calculated.
All filters and the stages of the Cascade Impactor were store in sealed bottles after weighing prior to subsequent analysis of captan concentration. Test atmospheres were sampled at least 3 times during each exposure period. The amount of material collectes on the filters was insufficient to permit accurate weighing for the 0.1 and 0.5 µg/l captan groups. Therefore only the 5 and 15 µg/L captan concentrations were monitored gravimetrlcally during exposure using pre- and post sampling filter weights. For target concentrations of 0.1 and 0.5 µg/L capatn, a SIMSLIN (Safety in Mines Scattered Light Instrument), connected to a pen recorder was used as an aerosol monitor. This gives a continuous analogue recording which Is related to particulate concentration, thus permitt ng control of the concentration of the atmosphere during exposure. The SIMSLIN was alternated between the 0.1 and 0.5 µg/L captan exposure chambers throughout the exposure period. The 15 µg/L captan group was also monitored using SIMSLIN, starting in week 5. For the reasons given above concerning accuracy of gravimetric analysis of the atmospheres, only the analysed concentrations of captan are reported. The aerodynamJc particle size of the test atmospheres were measured each day during the first week and once a week thereafter by means of a Marple Cascade Impactor {supplied by Shaeffer Instruments Ltd, Wantage, Oxon, UK) which aerodynainically separates airborne particles into pre-determined size ranges The mean amount of analysed captan, in each size range, was then used to calculate the aerodynamic particle size distribution of the aerosol. Using a nicrocomputer the data was transformed using a log/probit transform and a linear regression derived from the cumulative data. Using this regression line, the mass median aerodynamic diameter {D50) and geometric standard deviation (GSD) were calculated.

Duration of treatment / exposure:

13-week inhalation

Frequency of treatment:

six hours per day, 5 days per week

Doses / concentrations

Doses / concentrations 1

Dose / conc.:

0 mg/L air (nominal)

Remarks:

Doses / Concentrations:0.1 μg/L captan (10/sex/dose)Basis:nominal conc.

Doses / concentrations 2

Dose / conc.:

0.001 mg/L air (nominal)

Remarks:

Doses / Concentrations:0.5 μg/L captan (10/sex/dose)Basis:nominal conc.

Doses / concentrations 3

Dose / conc.:

0.005 mg/L air (nominal)

Remarks:

Doses / Concentrations:5.0 μg/L captan (10/sex/dose)Basis:nominal conc.

Doses / concentrations 4

Dose / conc.:

0.015 mg/L air (nominal)

Remarks:

Doses / Concentrations:15 μg/L captan (20/sex/dose)Basis:nominal conc.

No. of animals per sex per dose:

1. 0.1 μg/L captan (10/sex/dose)
2. 0.5 μg/L captan (10/sex/dose)
3. 5.0 μg/L captan (10/sex/dose)
4. 15 μg/L captan (20/sex/dose)5. control group of 20 males and 20 females was exposed to air only

Control animals:

yes, concurrent vehicle

Details on study design:

In a 13-week inhalation (nose only) study with captan (purity 88.7%, batch number 11240-37-1), ALpk:APfSD (Wistar derived) rats were exposed to nominal concentrations of 0.1, 0.5 and 5.0 μg/L captan (10/sex/dose) and 15 μg/L captan (20/sex) for six hours per day, 5 days per week. Test atmospheres were generated by the suspension of captan particulate in air using a Wright dust feed generator. A concurrent control group of 20 males and 20 females was exposed to air only.

Examinations

Observations and examinations performed and frequency:

Parameters evaluated were mortality and clinical signs (daily and frequently during exposure body weight and food consumption (weekly), ophthalmoscopy (pretest, week 13 (0 and 15 μg/L) and week 18 (‘recovery’ group), haematology and clinical chemistry (study termination), urinalysis (males in week 13 and females in week 14), gross pathology, organ weights (lungs (with trachea attached but larynx removed), heart, liver, kidneys, brain, adrenal glands and testes) and histopathology. Kidney sections from control and high dose animals were taken for immunocytochemical detection of α-2u-globulin.

Sacrifice and pathology:

At the end of the exposure period 10 males and 10 females from each group were sacrificed from the main study group. The remaining animals (10/sex/dose - control and 15 μg/L) were sacrificed after a 4-week ‘recovery’ period and are referred to as the satellite group.

Results and discussion

Results of examinations

Clinical signs:

effects observed, treatment-related

Description (incidence and severity):

Four males exposed to 12.98 μg/L captan were found dead over weeks 5 to 13 and another male from the same dose group was killed in extremis in week 11.

Mortality:

mortality observed, treatment-related

Description (incidence):

Four males exposed to 12.98 μg/L captan were found dead over weeks 5 to 13 and another male from the same dose group was killed in extremis in week 11.

Body weight and weight changes:

effects observed, treatment-related

Description (incidence and severity):

A statistically significant reduction in the body weight of males from all test groups compared to the control was noted during week 1. This divergence continued for the duration of the study for all test groups except 0.60 mg/L.

Food consumption and compound intake (if feeding study):

no effects observed

Description (incidence and severity):

Food intake in 5.06 and 12.98 μg/L males from the main group was reduced by approximately 10% versus controls, although these effects were not consistent and showed no dose relationship.

Food efficiency:

no effects observed

Description (incidence and severity):

Food efficiency was not affected by treatment.

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

not specified

Ophthalmological findings:

no effects observed

Haematological findings:

no effects observed

Clinical biochemistry findings:

effects observed, non-treatment-related

Description (incidence and severity):

Blood: There were several statistically significant differences between control and test groups however, these were small and were not dose-related and were therefore considered not to be compound-related.

Urinalysis findings:

no effects observed

Organ weight findings including organ / body weight ratios:

no effects observed

Gross pathological findings:

effects observed, non-treatment-related

Description (incidence and severity):

Gross findings in the lungs of animals dead or sacrificed prior to study termination (red discolouration and partial deflation) were considered to be agonal and not treatment-related.

Histopathological findings: non-neoplastic:

effects observed, treatment-related

Description (incidence and severity):

Microscopic changes related to inhalation of captan were seen in the lung, larynx, nasal passages and trachea.

Details on results:

General observations: Four males exposed to 12.98 μg/L captan were found dead over weeks 5 to 13 and another male from the same dose group was killed in extremis in week 11. One 0.13 μg/L female and one 5.06 μg/L female were killed in extremis in week 6 and week 10, respectively, but were considered to be not treatment-related. Any observed clinical signs were associated with restraint and occurred equally in test animals and controls. Respiratory noise was present towards the end of the study, primarily in those animals in the higher dose groups. Mucus nasal secretion was noted in a few females and may have been related to treatment. Body weight gain in females was similar in all dose groups throughout the course of the study. A statistically significant reduction in the body weight of males from all test groups compared to the control was noted during week 1. This divergence continued for the duration of the study for all test groups except 0.60 mg/L, which returned to control levels from week 2 onward. By week 13 final body weights were statistically decreased by 8, 7 and 8% compared to the control in the 0.13, 5.06 and 12.98 μg/L dose groups, respectively. These were not considered to be a toxicologically significant effect. The body weights of males in the main study are summarised in Figure 5.3.3.1-1 (look at attached background material).
In the satellite study, body weight gain by 12.98 μg/L males was significantly lower than controls from week 1 through to the end of the recovery period at week 17. Final body weight was significantly lower than the control (Figure 5.3.3.1-1), (look at attached background material).
Food intake in 5.06 and 12.98 μg/L males from the main group was reduced by approximately 10% versus controls, although these effects were not consistent and showed no dose relationship. Food efficiency was not affected by treatment.
Ophthalmoscopy haematology and clinical chemistry: No treatment-related effects were observed.
Gross pathology, organ weights and histopathology: There were no macroscopic changesrelated to treatment with captan. Gross findings in the lungs of animals dead or sacrificed prior to study termination (red discolouration and partial deflation) were considered to be agonal and not treatment-related.
Microscopic changes related to inhalation of captan were seen in the lung, larynx, nasal passages and trachea. Treatment related (and agonal) effects for intercurrent and main group animals are summarised in Table 5.3.3.1-2 (look at attached background material) and Table 5.3.3.1-3 (look at attached background material).
Following a 4-week recovery period the lung and nasal passage effects had resolved, butthe larengyl effects (Table 5.3.3.1-4) (look at attached background material) were still present in the high dose group.There was no evidence of any treatment-related increase in α-2u-globulin in the kidney in either males or females from the 12.98 μg/L dose group. There were no treatment-related effects on absolute or relative organ weights. The factor contributing to death in intercurrent males exposed to 12.98 μg/L captan was considered to be necrosis of the bronchial/bronchiolar epithelium. This effect was also seen in 5.06 and 12.98 μg/L captan animals that survived to terminal sacrifice. Other treatmentrelated effects were consistent with repeated exposure to an irritant particulate and areconsidered to have no toxicological significance in the context of captan exposure to humans.

Effect levels

Key result

Dose descriptor:

NOEC

Effect level:

0.6 mg/m³ air (nominal)

Based on:

test mat.

Sex:

male/female

Basis for effect level:

other: The NOEC for toxicological effects was 0.60 μg/L captan in both male and female rats, based on effects in the lung at doses of 5.06 μg/L and above.

Target system / organ toxicity

Critical effects observed:

not specified

Applicant's summary and conclusion

Conclusions:

The NOEC for toxicological effects was 0.60 μg/L captan in both male and female rats, based on effects in the lung at doses of 5.06 μg/L and above. It was not possible to establish an unequivocal NOEC.

Executive summary:

Inhalation of captan may form a route of human exposure. The purpose of this study was to assess the sub-chronic inhalation toxicity of captan, following exposure to rats for 13 weeks (5 exposures/week) to determine effect and no-effect levels. Satellite groups were attached to the control and highest exposure groups and were retained after the last exposure for a 4-week observation period.

The rat was chosen since there is considerable general and toxicologlcal data available to assist in the interpretation of results. The Alpk:APfSD strain was used in the study since relevant background data were available. Groups of 10 male and 10 female AlpK:APfSD rats were exposed nose-only to target concentrations of 0.1, 0.5 or 5.0 µg/l captan, and 20 male and 20 female rats were exposed to 15.0 µg/l captan, for 6 hours per day, 5 days per week for 13 weeks. A concurrent control group of 20 male and 20 female rats was similarly treated but was exposed to air only. Ten males and 10 females from each group were killed in week 14 following their last exposure, and the remaining animals were killed in week 18 following a 4-week 'recovery' phase.

The mean atmospheric concentrations of captan throughout the study, analysed by gas chromatography were 0.13, 0.60, 5.06 and 12.98 µg/l. The test atmospheres had study mean mass median aerodynamic diameters of 0.95, 1.22, 1.57 and l.60 µm respectively for the 0.13, 0.60, 5.06 and 12.98 µg/l captan exposure levels and respective geometric standard deviations of 1.82, 1.80, 1.84 and 2.00.

No treatment-related changes were seen in ophthalmoscopy, clinical chemistry or haematological parameters in any exposed group. No evidence of kidney toxicity was seen in this study. Treatment-related effects were confined to the respiratory tract, were consistent with exposure to an irritant particulate and affects in the lung were considered responsible for five male mortalities during the study in the group exposed to 12.98 µg/l captan. Lungs from animals allowed to recover for 4weeks were completely normal. 0.60 µg/l captan is considered to be a no-effect level for the lung. The larynx was the only other organ to be affected. Effects considered to be of toxicological significance were seen at 12.98 and 5.06 µg/l. Effects considered to be an adaptive response to irritants were seen at the lower concentration. The toxicological no-effect level for inhalation (NOEL) for captan was considered to be 0.60 µg/l.