Reaction mass of 2,2'-(ethylenedioxy)diethanol and 3,6,9,12,15-pentaoxaheptadecane-1,17-diol and 3,6,9,12-tetraoxatetradecane-1,14-d

Repeated dose toxicity: inhalation

Administrative data

Endpoint:

short-term repeated dose toxicity: inhalation

Type of information:

read-across based on grouping of substances (category approach)

Adequacy of study:

key study

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

study well documented, meets generally accepted scientific principles, acceptable for assessment

Justification for type of information:

Read across is based on the category approach. Please refer to attached category document.

Data source

Reference

Reference 1

Reference Type:

study report

Title:

Unnamed

Year:

1994

Report date:

1994

Reference 2

Reference Type:

publication

Title:

Respiratory peripheral chemosensory irritation, acute and repeated exposure toxicity studies with aerosols of triethylene glycol

Author:

B Ballantyne, WM Snellings, JC Norris

Year:

2006

Bibliographic source:

J Appl Toxicol 26:387-396

Materials and methods

Principles of method if other than guideline:

Nose only inhalation toxicity study.

GLP compliance:

yes

Limit test:

no

Test material

Test material information

Constituent 1

Reference substance name:

2,2'-(ethylenedioxy)diethanol

EC Number:

203-953-2

EC Name:

2,2'-(ethylenedioxy)diethanol

Cas Number:

112-27-6

Molecular formula:

C6H14O4

IUPAC Name:

2,2'-[ethane-1,2-diylbis(oxy)]diethanol

Specific details on test material used for the study:

Triethylene glycol, TEG
CAS 112-27-6
Clear liquid
Stored at room temperature
Purity: 99.9%

Test animals

Species:

rat

Strain:

other: CD

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Charles River Laboratories, Portage, MI
- Females (if applicable) nulliparous and non-pregnant: yes
- Age at study initiation: 8 weeks
- Weight at study initiation: 257.8-332 g (males), 168.5-242 g (females)
- Fasting period before study: Food available ad libitum except during exposures
- Housing: Individually in stainles stell wire mesh cages
- Diet (e.g. ad libitum): Ground Lab Diet The Richmond Standard Certified Rodent Diet #5002 (PMI Feeds, Inc.) available ad libitum except during exposures
- Water (e.g. ad libitum): Municipal tap water available ad libitum
- Acclimation period: 3 weeks

DETAILS OF FOOD AND WATER QUALITY: Water analyzed at regular intervals. EPA standards for maximum levels of contaminants were not exceeded. Analyses for chemical composition and possible contaminants of each feed lot were performed by PMI Feeds, Inc.

ENVIRONMENTAL CONDITIONS
- Temperature (°F): 66-77 degrees F
- Humidity (%): 40-70
- Air changes (per hr): Not provided
- Photoperiod (hrs dark / hrs light): 12/12

IN-LIFE DATES: From: To: Animals arrived on November 15, 1993 at 35 days old. Exposures began on December 6, 1993. Ten animals/sex/group were sacrificed on December 17, 1993. Five animals/sex of the control and high concentration groups assigned to the recovery group were sacrificed on January 14, 1994.

Administration / exposure

Route of administration:

inhalation: aerosol

Type of inhalation exposure:

nose only

Vehicle:

air

Mass median aerodynamic diameter (MMAD):

>= 1.2 - <= 1.4 µm

Geometric standard deviation (GSD):

1.32

Remarks on MMAD:

1.2, 1.4, and 1.3 microns for the 100, 500 and 1000 mg/m3 groups, respectively
GSD: 1.25-1.39

Details on inhalation exposure:

A polyvinyl chloride (PVC) exposure chamber was used. These exposure chambers utilize the flow-past, nonrebreathing concept (Cannon et al., 1983) for TEG exposure of the animals. The exposure chambers were composed of separate tiers, with each tier containing a total of 8 exposure ports for exposing 8 animals (1 exposure port/animal). Each animal was housed in a plexiglas tube (5.7 cm diameter tapered front x 19.5 cm length). The control and 1000 mg/m3 exposure chambers had 4 tiers, and the 100 and 500 mg/m3 exposure chambers had 3 tiers. Chambers were provided with air at a flowrate of approximately 16 lpm for the 0 and 1000 mg/m3 and 12 lpm for the 100 and 500 mg/m3 to ensure an adequate oxygen content of at least 19%. The airflow rates were monitored continuously and recorded approximately every 30 minutes. All chambers were maintained at a slightly negative pressure.

Liquid TEG was aerosolized, for all exposure levels, by positioning a single barrel Laskin Aerosol Generator (Enviro-Air Tech, Inc., Goshen, NY) into a glass 3-neck flask containing the TEG. The TEG aerosol was introduced to thenose-only exposure chamber by filtered compressed air passing through the nebulizer and flask and exhausting through 1-inch glass tubing connected to the inlet tube of the chamber. Operating air pressures for the nebulizers ranged from 4 to 6.25 psi.

Analytical verification of doses or concentrations:

yes

Details on analytical verification of doses or concentrations:

Chamber concentrations in each exposure chamber were determined 3 times during each exposure period by standard gravimetric techniques.
Mean gravimetric concentrations were 102 (+/- 8.2), 517 (+/- 37.9), and 1036 (+/- 27.2) mg/m3 for the target concentrations of 100, 500, and 1000 mg/m3 respectively.

The particle size distribution was measured using a TSI Aerodynamic Particle Sizer (TSI Incorporated, St. Paul, MN) and was determined each day for all exposure groups. The data collected were analyzed by probit analysis (Hinds, 1982) to obtain the mass median aerodynamic diameter (MMAD) and the geometric standard deviation (deltag).

Duration of treatment / exposure:

6 hours/day

Frequency of treatment:

5 consecutive days, 2 days without exposure (weekend), then 4 additional consecutive days

Doses / concentrations

Doses / concentrations 1

Dose / conc.:

102 mg/m³ air (analytical)

Remarks:

100 mg/m3 (targeted)

Doses / concentrations 2

Dose / conc.:

517 mg/m³ air (analytical)

Remarks:

500 mg/m3 (targeted)

Doses / concentrations 3

Dose / conc.:

1 036 mg/m³ air (analytical)

Remarks:

1000 mg/m3 (targeted)

No. of animals per sex per dose:

10/sex/dose (day 12 sacrifice)
5/sex/dose - high dose and control only (4 week recovery sacrifice)

Control animals:

yes, concurrent vehicle

Details on study design:

Animals were exposed for 6 hours/day for 5 consecutive days. The 6-hour exposure period was defined as the time when the animals were connected to and then disconnected from the exposure chamber. After 2 days without exposure (weekend), the animals were exposed for an additional 4 consecutive days. All control animals were exposed only to filtered air using the same exposure regimen. Ten animals/sex/group were sacrificed on December 17, 1993, the day after 9 exposures. The 5 animals/sex of the control and high concentration groups assigned to the recovery group were sacrificed on January 14, 1994, 4 weeks after the final exposure.

Examinations

Observations and examinations performed and frequency:

All animals were individually observed for signs of toxic effects, except during the exposures. During the exposures, observations were recorded on a group basis. Preceding and following each exposure, observations were recorded for animals exhibiting overt clinical signs. At the time of body weight collection and just preceding sacrifice, detailed observations were performed on all animals. On nonexposure days, the animals were observed once a day for overt clinical signs and twice a day for mortality. Body weights were measured for all animals on the morning prior to initiation of the first exposure (Study Day 1), Study Days 2, 5, 8, 9, and immediately preceding sacrifice. The animals were weighed weekly during the 4-week recovery period. Body weight gains were calculated for the periods between weighings. Food and water consumption were measured over Study Days 1-2, 2-5, 5-8, 8-10 (males) and 8-11 (females). Prior to the first exposure, the eyes of all rats were examined by a veterinarian using direct ophthalmoscopy following dilation of eyes with MYDRIACYL 1% (tropicamide 1.0%) Ophthalmic Solution. Following the eighth exposure for the female rats and following the ninth exposure for the male rats, the eyes were again examined by a veterinarian by direct ophthalmoscopy
following dilation of eyes. An ophthalmology examination was not performed on animals held for the 4-week recovery period.

Sacrifice and pathology:

Prior to the final sacrifice, blood was obtained from all surviving animals, with the exception of those designated for the recovery phase, for hematology and clinical chemistry determinations. Blood samples were collected by retroorbital bleeding from methoxyflurane anesthetized rats on Day 12. Rats were not fasted prior to bleeding. Feed was removed from all animal cages prior to the start of the blood collection period, but water was supplied ad libitum. Blood samples were not collected on the recovery animals. Following the eighth exposure (Study Day 10) for male rats and following the ninth exposure (Study Day 11) for female rats, urine was collected over an approximate 18-hour period from all rats, except those designated for the recovery phase. The rats were housed individually in Nalgene metabolic cages with stainless steel, wire mesh bottoms, approximately 22 cm diameter x 12 cm high (Nalge Company, Rochester, NY), during the collection period. Food and water were available ad libitum. Thymol crystals were added to the collection tube as a preservative. Urine samples were not collected on the recovery animals.

The following were measured or calculated:
Hematology: hematocrit total leukocyte count, hemoglobin differential leukocyte count, erythrocyte count platelet count, mean corpuscular volume (MCV) reticulocyte count, mean corpuscular hemoglobin (MCH), mean corpuscular hemoglobin, concentration (MCHC)

Clinical Chemistry: glucose (nonfasting), urea nitrogen, creatinine, total protein, protein electrophoresis, albumin, globulins, total bilirubin, direct bilirubin, indirect bilirubin (calculated), calcium, phosphorus, sodium, potassium, chloride, aspartate aminotransferase (AST), alanine aminotransferase (ALT), creatine kinase (CK), lactate dehydrogenase (LDH), gamma-glutamyl transferase (GGT), sorbitol dehydrogenase (SDH), alkaline phosphatase (ALK)

Urinalysis and Urine Chemistry: osmolality, PH, protein, glucose, ketones, bilirubin, creatinine, creatinine clearance (calculated), blood, urobilinogen, total volume, color and appearance, microscopic elements, N-acetyl-beta-D-glucosaminidase (NAG), alpha2U-globulin

At the end of the exposure regimen, all surviving animals, with the exception of those designated for the recovery phase, were anesthetized with halothane and were euthanized by severing the brachial vessels to permit exsanguination. On the day of sacrifice, body weight was obtained to allow expression of relative organ weights. A complete necropsy was performed on all sacrificed animals. The liver, kidneys, brain, adrenals, lungs, spleen and testes were weighed for all sacrificed animals. An additional 5 rats/sex/group from the control and high exposure groups were euthanized in the same manner following a 4-week recovery period. The recovery animals also received a complete necropsy. The following tissues were collected and retained in 10% neutral buffered formalin: gross lesions, lungs, nasopharyngeal tissue, brain, thymic region, trachea, heart, liver, spleen, kidneys, adrenals, testes, ovaries, urinary bladder, lymph nodes, mesenteric and non-mesenteric, nerve, sciatic, eyes, larynx. Gross lesions, lungs, naspharyngeal tissue, trachea, liver, kidneys, urinary bladder, and layrnx from all animals of the control and high exposure groups from the Day 12 sacrifice were processed histologically and examined microscopically. In addition, the lungs, liver, kidneys, and all gross lesions for all animals from the low and intermediate exposure groups were examined.

Statistics:

The data for quantitative continuous variables were intercompared for the 3 exposure groups and the control group by use of Levene's test for equality ofvariances, analysis of variance (ANOVA), and t-tests. The t-tests were used when the F value from the ANOVA was significant or only one group was compared to the control group. When Levene's test indicated similar variances, and the ANOVA was significant, a pooled t-test was used for pairwise comparisons. When Levene's test indicated heterogeneous variances, all groups were compared by an ANOVA for unequal variances followed, when necessary, by a separate variance t-test for pairwise comparisons. Nonparametric data were statistically evaluated using the Kruskal-Wallis test followed by the Mann-Whitney test. Incidence data were compared using Fisher's Exact Test. For all statistical tests, the probability value of < 0.05 (two-tailed) was used as the critical level of significance. Various models of calculators, computers, and computer programs may have been used to analyze data for this study. Since various models round or truncate numbers differently, values in some tables may differ slightly from those in other tables or from independently calculated data. The integrity of the study and interpretation of the data were unaffected by these differences.

Results and discussion

Results of examinations

Clinical signs:

no effects observed

Mortality:

mortality observed, non-treatment-related

Description (incidence):

One male in the control group died during the first exposure and was replaced with another animal. One male and 1 female were found dead in the 500 mg/m3 group on Days 1 and 2, respectively. These deaths were not considered related to the TEG exposure

Body weight and weight changes:

no effects observed

Description (incidence and severity):

By the end of the exposure period (day 12), both male and female rats of the 1036 mg/m3 group had body weight gains that were numerically lower (circa 15%) in comparison with the controls. In this group, the absolute body weights of males were similar to the controls throughout the study and the absolute weights of females were only slightly lower (4%) than the controls by the end of the study.

Food consumption and compound intake (if feeding study):

no effects observed

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

no effects observed

Ophthalmological findings:

no effects observed

Description (incidence and severity):

The only ophthalmic lesion found in any TEG exposed animals was mild unilateral conjunctivitis in 1 female rat from the 500 mg/m3 group. As no rats were affected from the high concentration group, this is considered to be an incidental finding. Animals saved for a recovery period were not reexamined as they had no eye lesions at the postexposure examination.

Haematological findings:

no effects observed

Clinical biochemistry findings:

effects observed, non-treatment-related

Description (incidence and severity):

Mean glucose levels were statistically significantly reduced in high dose females. This was not considered related to treatment.

Urinalysis findings:

no effects observed

Organ weight findings including organ / body weight ratios:

effects observed, non-treatment-related

Description (incidence and severity):

At the Day 12 sacrifice, no organ weight changes in the males or females were observed. At the Day 40 sacrifice, the absolute and relative (as percentages of body and brain weights) adrenal gland weights were increased in the males in the 1000 mg/m3 group. The relative (as a percentage of body weight) adrenal gland weights in the females increased in the 1000 mg/m3 group. As the organ weight changes occurred 4 weeks after the TEG exposures were terminated, they were not considered related to TEG.

Gross pathological findings:

no effects observed

Histopathological findings: non-neoplastic:

no effects observed

Histopathological findings: neoplastic:

no effects observed

Effect levels

Dose descriptor:

NOAEC

Effect level:

1 036 mg/m³ air (analytical)

Based on:

test mat.

Sex:

male/female

Basis for effect level:

other: No statistically significant, treatment-related changes at the high dose.

Target system / organ toxicity

Critical effects observed:

no

Applicant's summary and conclusion

Conclusions:

A 9 day repeated aerosol study was conducted, but by nose-only exposure of rats for 6 h/day to TEG aerosol concentrations of 0, 102, 517 and 1036 mg/m3. In this study there were no clinical signs, no effects on food and water consumption, and no biochemical or histological evidence of hepatorenal dysfunction. By the end of the exposure period, male and female rats of the 1036 mg/m3 group had body weights lower than those of the controls, but not with statistical significance. Since there were no statistically significant effects on any monitors, 1036 mg/m3 is considered to be a threshold for toxicity by nose-only exposure to TEG aerosol.

Executive summary:

A 9 day repeated aerosol study was conducted, but by nose-only exposure of rats for 6 h/day to TEG aerosol concentrations of 0, 102, 517 and 1036 mg/m3. In this study there were no clinical signs, no effects on food and water consumption, and no biochemical or histological evidence of hepatorenal dysfunction. By the end of the exposure period, male and female rats of the 1036 mg/m3 group had body weights lower than those of the controls, but not with statistical significance. Since there were no statistically significant effects on any monitors, 1036 mg/m3 is considered to be a threshold for toxicity by nose-only exposure to TEG aerosol.