N-(2-ethylhexyl)-8,9,10-trinorborn-5-ene-2,3-dicarboximide

Repeated dose toxicity: inhalation

Administrative data

Endpoint:

sub-chronic toxicity: inhalation

Type of information:

experimental study

Adequacy of study:

key study

Study period:

March - September 1993

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

1994

Report date:

1994

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

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

GLP compliance:

yes

Test material

Test material information

Constituent 1

Reference substance name:

N-(2-ethylhexyl)-8,9,10-trinorborn-5-ene-2,3-dicarboximide

EC Number:

204-029-1

EC Name:

N-(2-ethylhexyl)-8,9,10-trinorborn-5-ene-2,3-dicarboximide

Cas Number:

113-48-4

Molecular formula:

C17H25NO2

IUPAC Name:

N-(2-ethylhexyl)-8,9,10-trinorborn-5-ene-2,3-dicarboximide

impurity 1

Reference substance name:

(E)-1,4-Bis(2-ethylhexylamino)-2-butene-1,4-dione

Molecular formula:

C20H38N2O2

IUPAC Name:

(E)-1,4-Bis(2-ethylhexylamino)-2-butene-1,4-dione

impurity 2

Reference substance name:

1-(2-ethylhexyl)-1H-pyrrole-2,5-dione

Molecular formula:

C12H19NO2

IUPAC Name:

1-(2-ethylhexyl)-1H-pyrrole-2,5-dione

Test material form:

liquid

Details on test material:

Purity and characterisation analysis conducted on the following sample; Supplier: McLaughlin Gormley King ; Batch Number: AB9500

Specific details on test material used for the study:

Test Material: MGK® 264
Supplier: Mclaughlin Gormley King Company
8810 Tenth Avenue North
Minneapolis, Minnesota 55427
Date Received: 30 September 1991 and 21 January 1993
Lot No.: 3843
Active Ingredient: 98% MGK® 264 Insecticice Synergist
Description: Liquid
Expiration Date: None. Test material Was shown to be stable for the duration of the study.

Test animals

Species:

rat

Strain:

Sprague-Dawley

Details on species / strain selection:

Sprague-Dawley - derived (CD®) [CD® - Crl: (CD®)BR]
Supplier: Charles River Breeding Laboratories, Inc.Kingston, New York 124E4

Sex:

male/female

Details on test animals or test system and environmental conditions:

age at receipt - 6 weeks
age at initiation - 8 weeks

Non exposure periods: Animals were doubly housed in suspended stainless steel wire mesh cages during the first week of the acclimation period and individually house thereafter
Food was ad libitum; Purina Ceritified Rodent Chow® #5002 meal, Purina Mil's Inc., St. Louis,MO. Fresh food presenied as required.
water was ad libitum; by automated watering system (Elizabethtown Water Campany).
Approximately 12 hour · light/dark cycle (7 AM to 7 PM) via automatic timer. Temperature and relative humidity were monitored and recorded twice daily and
maintained, to the maximum extent possible, within the range presented below.
Temp: Desired: 20-24°C
Actual : 22 ± 1 °C (X ± S. D.)
Humidity: Desired: 40-70%
Actual: 54 ± 9% (X ± S.D.)

During Exposure: Animals were individually housed in stainless steel, wire mesh cages within a 1L glass and stainless steel exposure chamber. no water/food
Temp: Desired: 20-24°C
Actual : 24 ± 1 °C (X ± S. D.)
Humidity: Desired: 40-60%
Actual: 50 ± 1% (X ± S.D.)

Administration / exposure

Route of administration:

inhalation: aerosol

Type of inhalation exposure:

whole body

Vehicle:

air

Mass median aerodynamic diameter (MMAD):

1.1 µm

Geometric standard deviation (GSD):

2.3

Remarks on MMAD:

It was concluded that the liquid aerosol was highly respirable to the rat with a similar MGK 264 particle size amongst all the test material exposed groups.

Details on inhalation exposure:

Appropriate amounts of MGK® 264 were placed into a 3-neck flask resting in a heating mantle regulated by a variable autotransformer. House-line air was delivered through a regulator and backpressure gauge via 1/4" plastic tubing to a plastic T-tube where it split the airflow to the generation and dilution systems. For the generation system the air was directed from the T - tube, to a Nupro® metering valve, Dwyer® flowmeter and Matheson® backpressure gauge. The air was then carried to a 1 or 2-barrel Laskin nebulizer contained inside a 3-neck flask. For the dilution system, air was directed d to a Nupro® metering valve and Dwyer® flowmeter where it converged at a glass T-Tube with the generation air to generate the test atmosphere. The resultant liquid aerosol was directed into the chamber which housed the animals. The animals remained in the chamber for 30 minutes following the exposure to allow the chamber to clear, using room air at the same airflow rate used during exposure.

Analytical verification of doses or concentrations:

yes

Remarks:

Gravimentrical determinations were made 4 times per chamber per day and gas chromatatography was performed once per chamber per day.

Details on analytical verification of doses or concentrations:

The nominal concentration was determined by weighing the generation apparatus containing the test material before and after the exposure and dividing the difference in these weights by the total volume of air delivered during exposure (volumetric flow rate times total exposure time).

Duration of treatment / exposure:

6 hours per day
Exposure levels were determined gravimetrically (four times per chamber per day) and by Gas Chromatography (GC) (one time per chamber per day). Particle size distribution measurements of the liquid aerosol were made once per chamber/day using a Delron DCI-6 cascade impactor.

Frequency of treatment:

5 days per week, for 13 weeks

Doses / concentrations

Doses / concentrations 1

Dose / conc.:

0 mg/m³ air (nominal)

Remarks:

Group 1 and 2

Doses / concentrations 2

Dose / conc.:

10 mg/m³ air (nominal)

Remarks:

Group 3

Doses / concentrations 3

Dose / conc.:

40 mg/m³ air (nominal)

Remarks:

Group 4

Doses / concentrations 4

Dose / conc.:

135 mg/m³ air (nominal)

Remarks:

Group 5

Doses / concentrations 5

Dose / conc.:

400 mg/m³ air (nominal)

Remarks:

Group 6

Doses / concentrations 6

Dose / conc.:

400 mg/m³ air (nominal)

Remarks:

Group 7

No. of animals per sex per dose:

15/sex/group

Control animals:

yes, concurrent vehicle

Examinations

Observations and examinations performed and frequency:

Physical observations for abnormal signs were performed once during each exposure for all animals; detailed physical examinations were conducted on all animals once pretest and weekly thereafter. Ophthalmoscopic examinations were performed on all animals pretest and once prior to scheduled sacrifice. Body weight measurements were recorded twice pretest, weekly thereafter and just prior to scheduled sacrifice. Food consumption measurements were conducted once pretest and weekly thereafter. Blood samples for analysis of hematology and clinical chemistry parameters were withdrawn just prior to the scheduled sacrifices. Following 13 weeks of exposure, 7 (groups I and VI) or 8 (Groups II and VII) animals/sex/group were sacrificed. Recovery animals were allowed a 13 week recovery period prior to sacrifice. Selected organs were weighed and organ/body and organ/brain weight ratios calculated for all animals. Complete macroscopic postmortem examination of selected tissues were conducted on all animals.

Sacrifice and pathology:

Following 13 weeks of exposure, 7 (groups I and VI) or 8 (Groups II and VII) animals/sex/group were sacrificed. Recovery animals were allowed a 13 week recovery period prior to sacrifice.

Statistics:

Body weight, change in body weight from Week 0, food consumption, haematology and clinical chemistry parameters, organ weights, organ/body and organ/brain weight ratios were analyzed. Mean values of the two control groups (I and II) and mean values of the two high dose groups (VI and VII) were compared in order to show comparable results regarding test material effects. When combined, the mean values of all exposure groups were compared to control at each time interval.

Results and discussion

Results of examinations

Clinical signs:

effects observed, treatment-related

Description (incidence and severity):

In-Chamber Observations: The only clearly test material related observation recorded during the exposures was decreased activity. This was seen in almost all animals in the high exposure groups over the entire study.

Weekly Detailed Observations: Weekly detailed observations noted dose-related incidences of red facial stains, nasal discharge and excessive salivation in both sexes. Excessive salivation occurred almost exclusively at the high exposure level and primarily during the first two weeks of the study. The red facial stains and nasal discharge were present over the entire exposure period. Red nasal discharge or staining is a common finding in inhalation studies. The animals were essentially all within normal limits during the recovery period.

Mortality:

mortality observed, non-treatment-related

Description (incidence):

One control group male was found dead on Day 46 of the study. One high exposure level male was sacrificed moribund during the recovery period. Neither of these deaths is considered to be test material related.

Body weight and weight changes:

no effects observed

Food consumption and compound intake (if feeding study):

no effects observed

Food efficiency:

not examined

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

not examined

Ophthalmological findings:

no effects observed

Haematological findings:

no effects observed

Clinical biochemistry findings:

no effects observed

Urinalysis findings:

not examined

Behaviour (functional findings):

not examined

Immunological findings:

not examined

Organ weight findings including organ / body weight ratios:

no effects observed

Gross pathological findings:

no effects observed

Neuropathological findings:

not examined

Histopathological findings: non-neoplastic:

effects observed, treatment-related

Description (incidence and severity):

Microscopic morphologic abnormalities considered to be related to the whole-body exposure to MGK® 264 were seen in the nasoturbinal tissues, nasopharynx and in the larynx of both males and females. In the nasoturbinates, these changes were seen and included an increased severity of hypertrophy/hyperplasia of goblet cells in the respiratory epithelium of males and a slight increase in the severity of intracytoplasmic eosinophilic material in the epithelium covering the respiratory and/or olfactory mucosa in males and females. In the nasopharynx, an increased incidence of hypertrophy/hyperplasia of the goblet cells in the epithelium lining the nasopharynx was seen in the males and females of the high exposure group. In the larynx, an increased severity of subacute (chronic active)/chronic inflammation was seen in the high exposure group. Squamous/squamoid metaplasia/hyperplasia of the pseudostratified columnar epithelium, frequently keratinized, of the larynx was seen in all treated groups (almost all animals) but not in any control animals. Hyperplasia and hyperkeratosis of the stratified squamous epithelium was seen in groups receiving exposures of 135 or 400 mg/m3. These changes were considered to be a direct local effect of the test material. At the end of the month post exposure recovery period, all of these findings had for all practical purposes, completely reversed.

Histopathological findings: neoplastic:

no effects observed

Effect levels

Key result

Dose descriptor:

NOAEL

Effect level:

ca. 135 mg/m³ air (nominal)

Based on:

test mat.

Sex:

male/female

Basis for effect level:

clinical signs

histopathology: non-neoplastic

Target system / organ toxicity

Key result

Critical effects observed:

no

Lowest effective dose / conc.:

400 mg/m³ air (nominal)

System:

respiratory system: upper respiratory tract

Organ:

larynx

other: nasoturbinates and nasopharynx

Any other information on results incl. tables

Tabulated data are presented in the final report which is attached to this end point summary.

Applicant's summary and conclusion

Conclusions:

Exposures to MGK® 264 as a liquid aerosol for 6 hours/day, 5 days/week for 13 weeks at levels of 10, 40, 135 and 400 mg/m3, produced test material related increases in red facial stains and nasal discharge in all test material exposure groups and excessive salivation in the high exposure group. Decreased activity was seen in the high exposure group during the exposures.
Body weight gain, food consumption, ophthalmology, hematology and clinical chemistry parameters, organ weights and organ/body weight ratios were not affected by these exposures.
Microscopic findings related to the test material included reversible changes in the nasoturbinates, nasopharynx and larynx.
All of these microscopic results were considered to be localized, normal adaptive responses to an aerosol exposure, and not indicative of systemic toxicity. Therefore, 135 mg/m3 was considered to be a NOAEL No-observable-adverse-effect-level).

Executive summary:

The toxicity of MGK 264 was assessed when administered by whole-body inhalation as a liquid aerosol to rats for 6 hours per day, 5 days per week for 13 weeks. The target concentrations in this study weere 0 (control; double group), 10, 40, 135 and 400 (double group) mg/m3. The highest concentration in the study represented the maximum exposure level of MGK 264 which could be generated at the appropriate particle size in the 1000 litre chambers that were used in the study. Recovery animals were included in the control and high exposure groups and were examined after a 13 week recovery period.

Exposure levels were determined gravimetrically (4 times per day) and by gas chromatography (once per day). Particle size distribution measurements of the liquid aerosol were made once per chamber/day.

The animals were observed for clinical signs during each exposure and weekly for detailed physical examinations. Body weight measurements were recorded pre-test and weekly. Ophthalmoscopy examinations were performed pre-test and at the end of the study. Blood samples wre withdrawn for haematological and clinical chemistry examinations before the scheduled sacrifices at the end of the treatment or recovery periods. At necropsy examination, selected organs were weighed, a full list of organs were harvested and processed for histopathological examination.

The mean active ingredient exposure concentrations were determined to be 0, 0, 10±2 (mean± standard deviation), 43±6, 135±19, 400±28 and 407±41 mg/m3 for

Groups I to VII, respectively. Particle size distribution determinations indicated the test aerosol atmosphere was respirable in size to the rat.

Particle size distribution determinations showed, on average, the mass median aerodynamic diameter to be 1.1 microns with a geometric standard deviation of

2.3.

MGK 264 related changes were limited to reduced activity and excessive salivation during exposure in the high exposure groups and histopathological changes in the nasoturbinates, nasopharynx and larynx of the high exposure group. These changes which were reversible, were indicative of a localised, normal adaptive responses to an aerosol exposure and not indicative of systemic toxicity. The decreased activity was not considered to be a major detrimental response. On this basis, 135 mg/m3 was considered to be a NOAEL (no-observable-adverse-effect-level).