ε-caprolactam

Repeated dose toxicity: inhalation

Administrative data

Endpoint:

sub-chronic toxicity: inhalation

Type of information:

experimental study

Adequacy of study:

key study

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Data source

Reference

Reference 1

Reference Type:

study report

Title:

Unnamed

Year:

1997

Report date:

1997

Reference 2

Reference Type:

publication

Title:

Subchronic Inhalation Toxicity Study of Caprolactam (with a 4-Week Recovery) in the Rat via Whole-Body Exposures.

Author:

Reinhold R.W. et al.

Year:

1988

Bibliographic source:

Toxicological Sciences 44, 197-205

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

other: described by TSCA, EPA September 1985 and May 1989

GLP compliance:

yes

Remarks:

(Huntington Life Science)

Limit test:

no

Test material

Test material information

Constituent 1

Reference substance name:

ε-caprolactam

EC Number:

203-313-2

EC Name:

ε-caprolactam

Cas Number:

105-60-2

Molecular formula:

C6H11NO

IUPAC Name:

azepan-2-one

Specific details on test material used for the study:

- Name of test material (as cited in study report): Caprolactam
- Physical state: white crystalline solid
- Analytical purity: 99.9%
- Stability under test conditions: stable for study duration
- Storage condition of test material: room temperature

Test animals

Species:

rat

Strain:

Sprague-Dawley

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Charles River Laboratories
- Age at study initiation: 6 weeks
- Mean weight at study initiation: males 178 g (151-205 g), females 143 g (110-167 g)
- Housing: 2 per cage
- Diet (e.g. ad libitum): Certified Rodent Diet, No. 5002, PMI feeds Inc, MO.
- Water: ad libitum
- Acclimation period: 2 weeks


ENVIRONMENTAL CONDITIONS
- Temperature (°C): 20-27
- Humidity (%): 21-74
- Photoperiod (hrs dark / hrs light): 12/12

Administration / exposure

Route of administration:

inhalation: aerosol

Type of inhalation exposure:

whole body

Vehicle:

water

Remarks on MMAD:

MMAD / GSD: 2.9 µm/1.7

Details on inhalation exposure:

GENERATION OF TEST ATMOSPHERE / CHAMBER DESCRIPTION
- Method of holding animals in test chamber: The placement of the animal in the whole body exposure chamber was rotated at each exposure to ensure uniform exposure of the animals.
- System of generating particulates/aerosols: A 1:1 solution, prepared by dissolving Caprolactam in an equal amount, by weight, of distilled demineralized water, was placed in a 250 ml Erlenmeyer flask and continuously stirred on a stirplate. The solution was fed directly, via an FMI Fluid Metering Pump, into the fluid inlet of a Spraying System, Inc. air atomizing nozzle (JCO-SS with a lA spray setup). The aerosol so generated was mixed with incoming house air by turbulence. Conditioned house air was metered into the control chamber.
- Temperature, humidity in air chamber: 21-32°C, 28-65%
- Method of particle size determination: Particle size distribution samples were withdrawn once during each exposure and determined using an aerodynamic particle sizer (TSI Aerodynamic Particle Sizer, Model 3300, and a Model 3302 TSI Aerosol Diluter). The mass median aerodynamic diameter (MMAD) and geometric standard deviation (GSD) were calculated.


TEST ATMOSPHERE
- Brief description of analytical method used: During each exposure, chamber concentration determinations were performed six times daily using gravimetric sampling. In addition, for each daily exposure group, one sample was also analyzed by high-pressure liquid chromatography (HPLC) using a Hewlett-Packard 1050 LC system and a UV detector.
- Samples taken from breathing zone: yes


VEHICLE (if applicable)
- Concentration of test material in vehicle: 1:1 aqueous solution with Caprolactam
- Lot/batch no. of vehicle (if required): Prepared by Pharmacy from water supplied by the Elizabethtown Water Company
- Purity of vehicle: Monthly analysis was provided by the Elizabethtown Water Company

Analytical verification of doses or concentrations:

yes

Details on analytical verification of doses or concentrations:

The mean (±SD) analytical exposure concentrations were 0±0, 23±4.9, 66±20 and 244±37 mg/m3 (approx. 0±0, 0.023±0.0049, 0.066±0.02 and 0.245 ±0.037 mg/l).
Gravimetrically determined exposure concentrations were (mean ±SD) 0.063 ±0.19, 24±5.6, 70±18 and 243±36 mg/m3 for the 13 weeks of exposure.

Duration of treatment / exposure:

13 weeks plus 4 week-recovery

Frequency of treatment:

6 hours/day, 5 days/week

Doses / concentrations

Doses / concentrations 1

Dose / conc.:

0 mg/L air (nominal)

Remarks:

0 mg/mL analytical conc.

Doses / concentrations 2

Dose / conc.:

0.025 mg/L air (nominal)

Remarks:

0.023 mg/mL analytical concentration

Doses / concentrations 3

Dose / conc.:

0.075 mg/L air (nominal)

Remarks:

0.066 mg/mL analytical concentration

Doses / concentrations 4

Dose / conc.:

0.25 mg/L air (nominal)

Remarks:

0.245 mg/mL analytical concentration

No. of animals per sex per dose:

10

Control animals:

yes

Details on study design:

- Post-exposure recovery period in satellite groups: An additional 10 animals/sex/group were similarly exposed and then held for a 4-week recovery period

Examinations

Observations and examinations performed and frequency:

CAGE SIDE OBSERVATIONS: Yes
- Time schedule: twice daily throughout the study and at least once during each exposure
- Cage side observations checked: mortality, general appearance and signs of toxic or pharmacological effects


BODY WEIGHT: Yes
- Time schedule for examinations: Twice pretest, weekly during study period and prior to the terminal and recovery sacrifices


FOOD CONSUMPTION:
- Food consumption for each animal determined: Yes


OPHTHALMOSCOPIC EXAMINATION: Yes
- Time schedule for examinations: prior to the first exposure and prior to the terminal sacrifice
- Dose groups that were examined: all


HAEMATOLOGY: Yes
- Time schedule for collection of blood: at the exposure and recovery terminations
- Anaesthetic used for blood collection: Yes (light carbon dioxide anesthesia)
- Animals fasted: Yes
- How many animals: all
- Parameters checked: erythrocyte count and morphology, total and differential leukocyte count, platelet count, hematocrit, hemoglobin concentration, mean corpuscular hemoglobin, mean corpuscular hemoglobin concentration, prothrombin time, and activated partial thromboplastin time.


CLINICAL CHEMISTRY: Yes
- Time schedule for collection of blood: at the exposure and recovery terminations
- Animals fasted: Yes
- How many animals: all
- Parameters checked: serum alanine aminotransferase, serum aspartate aminotransferase, alkaline phosphatase, blood urea nitrogen, creatinine, fasting glucose, total serum protein, albumin, globulin (calculated as total protein - albumin = globulin), albumin/globulin ratio (calculated), total bilirubin, sodium, potassium, chloride, calcium, inorganic phosphorus, and y-glutamyl transferase


NEUROBEHAVIOURAL EXAMINATION: Yes
- Time schedule for examinations: Pretest and Weeks 5, 9, and 14 and Recovery (Week 19)
- Dose groups that were examined: all
- Battery of functions tested: sensory activity / grip strength / motor activity / other: For homecage observations, each rat was observed and scored for posture, vocalization, and palpebral closure. On removal from the cage, each animal was observed and scored for ease of removal, ease of handling, lacrimation, appearance of fur, and salivation. Open-field evaluations included observations and scoring of gait, degree of locomotion, arousal, abnormal movements including tremors or convulsions, exophthalmia, piloerection, and presence or absence of diarrhea and urination. Reflex assessments consisted of scoring of responses to approach, tail pinch, finger snap, pupillary response, and air righting reflex. Additionally, landing foot splay and fore- and hindlimb grip strengths were assessed. Motor activity was monitored using an automated Photobeam Activity System (San Diego Instruments, Inc.). Sessions were 60 min in length; each session was divided into 12 intervals of 5-min duration. During each 5-min activity session, activity counts (photobeam breaks) were recorded at 1-min intervals. Treatment groups were counterbalanced across test times.

Sacrifice and pathology:

GROSS PATHOLOGY: Yes
HISTOPATHOLOGY: Yes

Other examinations:

At necropsy, the adrenals, brain, liver, lungs, kidneys, ovaries, and testes (with epididymides) were weighed and recorded and organ/body and organ/brain weight ratios were calculated.

Results and discussion

Results of examinations

Clinical signs:

effects observed, treatment-related

Description (incidence and severity):

Treatment- related responses such as labored breathing and nasal discharge were seen during many of the exposures at all exposure levels. A slight increase in incidence was seen with increasing exposure concentration. Similar responses as well as moist rales were seen during the non-exposure periods during the 13 weeks of exposure. These responses abated during the 4-week recovery period.

Mortality:

no mortality observed

Description (incidence):

All animals survived to the termination of the study.

Body weight and weight changes:

effects observed, non-treatment-related

Food consumption and compound intake (if feeding study):

effects observed, non-treatment-related

Food efficiency:

not examined

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

not examined

Ophthalmological findings:

effects observed, non-treatment-related

Haematological findings:

effects observed, non-treatment-related

Clinical biochemistry findings:

effects observed, non-treatment-related

Urinalysis findings:

not examined

Behaviour (functional findings):

effects observed, non-treatment-related

Immunological findings:

not examined

Organ weight findings including organ / body weight ratios:

effects observed, non-treatment-related

Gross pathological findings:

effects observed, non-treatment-related

Neuropathological findings:

effects observed, non-treatment-related

Histopathological findings: non-neoplastic:

effects observed, treatment-related

Description (incidence and severity):

Microscopic findings which were considered to be related to the test material were seen in the nasoturbinal tissues (hypertrophy/hyperplasia of goblet cells in the respiratory mucosa and intra-cytoplasmic eosinophilic material in epithelial cells of the olfactory mucosa) of the 2 higher exposure levels and in the larynx (squamous/ squamoid metaplasia/ hyperplasia of the pseudostratified columnar epithelium covering the ventral seromucous gland) of all exposure levels. Minimal keratinization of the metaplastic epithelium of the larynx was seen in a small number of the 245 mg/m3 exposed rats. Some recovery was seen in both nasoturbinal and laryngeal tissues following the 4-week recovery period, but the recovery was incomplete.

Effect levels

1

Dose descriptor:

NOAEC

Effect level:

0.066 mg/L air (analytical)

Sex:

male/female

Basis for effect level:

other: local effects: laryngeal keratinization

2

Dose descriptor:

NOAEC

Effect level:

0.245 mg/L air (analytical)

Sex:

male/female

Basis for effect level:

other: no systemic effects at highest dose level

Target system / organ toxicity

Critical effects observed:

not specified

Applicant's summary and conclusion

Executive summary:

A study descrobed by TSCA, EPA September 1985 and May 1989 was performed.

The whole-body exposure of Sprague-Dawley rats to Caprolactam as an aerosol for 6 hours/day, 5 days/week for 13 weeks at analytically determined levels of 0.023, 0.066 and 0.245 mg/l resulted in transient clinical signs (secretory and respiratory) as well as respiratory tract effects (microscopically-detected nasoturbinal and/or laryngeal tissue changes) at all exposure levels with incomplete recovery within 4 weeks after exposure. However, these effects, except the laryngeal keratinization at 0.245 mg/l, were believed to be a normal physiological response to inhaled foreign particulate material and were not believed to represent an adverse response.This interpretation is fully supported by the proceedings of an international expert workshop of the European Society of Toxicology (Kaufmann et al., 2009, Exp. Tox. Path., 61, 591-603):slight laryngeal squamous metaplasia that are not observed diffusely could occur spontaneously or as treatment-induced lesions and should be assessed as “non-adverse”. Therefore, under the conditions of this study, a NOAEC (No Observed Adverse effect Level) was considered to be 0.066 mg/l due to effects on the upper respiratory tract. However, under the conditions of this study, a NOAEC was considered to be 0.245 mg/l due to no effects on the lower respiratory tract, systemic toxicity and neurotoxicity.