Trifluoroiodomethane

Endpoint summary

Description of key information

Two sub-chronic and several sub-acute toxicity are available. Based on the key study a NOAEC of 20000 ppm (or 160254 mg/m3) is derived.

Key value for chemical safety assessment

Repeated dose toxicity: via oral route - systemic effects

Endpoint conclusion

Endpoint conclusion:

no study available

Repeated dose toxicity: inhalation - systemic effects

Link to relevant study records

Reference

Endpoint:

sub-chronic toxicity: inhalation

Type of information:

experimental study

Adequacy of study:

key study

Study period:

1996 to 1998

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

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

Reason / purpose for cross-reference:

reference to same study

Reason / purpose for cross-reference:

reference to same study

Reason / purpose for cross-reference:

reference to same study

Qualifier:

no guideline followed

Principles of method if other than guideline:

A combined repeated dose and reproductive toxicity screening study was performed in Sprague-Dawley Crl:CD(SD)BR rats. The study had 16 male and 16 female rats/group to yield at least 12 pregnant females at term. The animals were assigned to one of four groups (0, 0.2, 0.7, and 2.0%) and exposed to substance vapor for 4 wk (6 h/day, 5 days/wk) prior to mating. An additional six male and three female rats were assigned to a fifth group only to serve as positive controls in the micronuclei assay. Animals were exposed 6 h/day, 7 days/wk, during the mating, gestation, and lactation phases of the study. However, dams were not exposed from gestation day 21 through lactation day 4 to allow for parturition and early lactation. Pups were not exposed to and remained in their home cages under observation. Following the 14-day mating period, eight male rats/group were terminated (study wk 7). The remaining 8 male rats/group and 16 female rats/group were terminated over a 5-day period subsequent to when the last female rat on study reached lactation day 21. Animals were killed by CO2 inhalation overdose followed by exsanguination.
Examinations included clinical observations (twice daily), bodyweight, haematology, clinical chemistry, serum thyroid hormone levels and bone marrow erythrocyte micronucleus test (summarized in section 7.6.2). At termination a gross examination was performed, organ weights were determined and selected tissues from male and female rats of the control and high-dose groups were subjected to histopathologic examination. Reproductive indices and parameters were assessed for all dose groups.

GLP compliance:

no

Limit test:

no

Specific details on test material used for the study:

Supplier: Ajay North America, LLC, Powder Springs, GA
Purity > 99.78%
Appearance: Colourless gas

Species:

rat

Strain:

Sprague-Dawley

Remarks:

Crl:CD(SD)BR

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Charles River Laboratories, Wilmington, MA.
- Females (if applicable) nulliparous and non-pregnant: yes
- Age at study initiation: approximately 11 weeks at the start of exposure
- Weight at study initiation: around 225 g at SD 0. Body weights of males are not reported.
- Housing: The rats were housed in wire-bottom cages during exposure (one per cage except during the mating period) and in plastic shoebox cages with bedding (two males per cage, one female per cage, except during the mating period) during nonexposure periods. Exposure cages were rotated in a clockwise manner within the chamber each exposure day.
- Diet: certified rodent diet (PMI Feeds, Inc., St. Louis, MO) was available ad libitum during nonexposure periods.
- Water: water was available ad libitum during nonexposure periods.
- Acclimatization period: 2 to 3 weeks

ENVIRONMENTAL CONDITIONS
- Temperature (°C): between 21 and 26
- Humidity (%): between 35 and 65
- Photoperiod (hrs dark / hrs light): 12/12

Route of administration:

inhalation: vapour

Type of inhalation exposure:

whole body

Vehicle:

air

Details on inhalation exposure:

GENERATION OF TEST ATMOSPHERE / CHAMBER DESCRIPTION
- Exposure apparatus: Whole-body inhalation exposures were performed in 690-L chambers made of stainless steel and glass.
- Source and rate of air: The substance and air for dilution were controlled through flow meters
- Temperature, humidity, oxygen and ammonia in air chamber: Relative humidity and temperature of the exposure atmosphere were constantly monitored and recorded. Oxygen concentration of the high concentration chamber was analyzed on occasion using a Hudson O2 sensor model 82T (MDA, Lincolnshire, IL). Ammonia levels during exposure were estimated not to be of biological importance
- Air flow and air change rate: To conserve the use of test material and minimize cost, total chamber air flow was reduced to 60 L/min (approximately 5 air changes/h).

Analytical verification of doses or concentrations:

yes

Details on analytical verification of doses or concentrations:

Continuous analysis of the chamber air for test substance was performed using infrared absorption spectrometers. Instrumental calibration was performed using known concentrations of freshly prepared test substance in air contained in Tedlar sample bags. Calibration checks were performed at appropriate intervals.

Duration of treatment / exposure:

14 weeks

Frequency of treatment:

- 6 hours/day, 5 days/wk prior to mating
- 6 hours/day, 7 days/ wk during the mating, gestation, and lactation phases of the study.
- Dams were not exposed from gestation day 21 through lactation day 4 to allow for parturition and early lactation.
- Pups were not exposed

Dose / conc.:

2 000 ppm

Remarks:

equivalent to 0.2%

Dose / conc.:

7 000 ppm

Remarks:

equivalent to 0.7%

Dose / conc.:

20 000 ppm

Remarks:

equivalent to 2.0%

No. of animals per sex per dose:

16. An additional six male and three female rats were assigned to a fifth group only to serve as positive controls in the micronuclei assay (section 7.6.2).

Control animals:

yes, concurrent vehicle

Details on study design:

- Dose selection rationale: 2 and 13 wk repeated dose toxicity studies are available. Exposure concentrations selected for this study extended beyond those of the 13-wk study (Dodd 1997)

Positive control:

Single dose of cyclophosphamide (7.5 mg/kg) for the micronucleus assay (please refer to section 7.6.2).

Observations and examinations performed and frequency:

CAGE SIDE OBSERVATIONS
The animals were observed twice daily (morning and afternoon), including inhalation exposure periods. Signs of toxicity were recorded.

BODY WEIGHT
The body weight of the male rats were determined prior to the first exposure and weekly thereafter. The body weights of female rats were determined and recorded in the same manner until confirmation of mating. During gestation, female rats were weighed on gestational days 0, 7, 14, and 20. Dams producing litters were weighed on days 0, 4, 7, 12, and 21 postpartum.

HAEMATOLOGY
Routine haematology evaluations were conducted on blood samples taken immediately prior to termination from all animals. The blood was sampled via the posterior vena cava. Haematologic parameters were determined according to established procedures, utilizing a Cell-Dyn 3500 (Abbott Diagnostics, Chicago). Assessed parameters included blood cell count, haemoglobin, mean corpuscular volume, mean corpuscular haemoglobin, mean corpuscular haemoglobin concentration, hematocrit, total and differential leukocyte count and platelet count.

CLINICAL CHEMISTRY
Routine clinical chemistry evaluations were conducted on blood samples taken immediately prior to termination from all animals. The blood was sampled via the posterior vena cava. Serum chemistry parameters were determined according to established procedures, utilizing a Vitros 250XR (Johnson and Johnson, Rochester, NY). Assessed parameters included total protein, albumin, globulin, alanine aminotransaminase (ALT), aspartate aminotransaminase (AST), alkaline phosphatase, urea nitrogen, creatinine, calcium, glucose, potassium, phosphorus, sodium, magnesium, trigylcerides and cholesterol.

SERUM THYROID LEVELS
The following serum thyroid hormone levels were determined in control and exposed rats: thyroxine (T4), triiodothyronine (T3), reverse T3 (rT3), and thyroid-stimulating hormone (TSH).
Assays for T4, T3, rT3, and TSH were performed using radioimmunoassay (RIA) kits and were carried out according to manufacturer’s instructions. For all thyroid hormone or TSH measurements, assay kits were prepared for each animal termination period (study wk 7 or 14) with the same batch number and the same expiration date. Tracer (125I) radioactivity was measured with a Packard gamma counter (Packard Instrument Co., Meriden, CT). For T3, the RIA assay kit was purchased from Diagnostic Product Corp. (Los Angeles, CA), and canine T3 antibody-coated tubes were used. For T4, the RIA assay kit was purchased from Diagnostic Product Corp. (Los Angeles, CA), and T4 antibody coated tubes were used. For rT3, the RIA assay kit was purchased from Wein Laboratories (Succasunna, NJ), and rT3 antiserum raised in the rabbit was used. For TSH, the RIA assay kit was purchased from Amersham Corp. (Arlington Heights, IL), and both lyophilized rabbit anti-rat TSH serum and Amerlex-M second antibody (donkey anti-rabbit serum coated onto magnetized polymer particles containing sodium azide) were used.

Sacrifice and pathology:

SACRIFICE
Following the 14-day mating period, eight male rats/group were terminated (study wk 7). The remaining 8 male rats/group and 16 female rats/group were terminated over a 5-day period subsequent to when the last female rat on study reached lactation day 21. Animals were not fasted prior to termination, since fasting might affect thyroid hormone levels.

GROSS NECROPSY
A gross examination was performed at necropsy.

HISTOPATHOLOGY / ORGAN WEIGHTS
Organs weighed included adrenal glands, heart, lungs, liver, kidneys, ovaries, testes, epididymides, brain, spleen, and thymus. Thyroid glands (including parathyroid glands) were removed with the trachea, fixed in buffered formalin, then dissected free of the trachea, and weighed. A single technician performed the dissection of the thyroid glands. These organs and the pituitary gland, nasal cavity, trachea, bronchi, bone marrow (sternal and femoral sections and smear), pancreas, urinary bladder, stomach, duodenum, ileum, colon, prostate, scrotum, seminal vesicles, vagina, uterus, and other tissues with gross lesions were removed from all animals and preserved in 10% neutral-buffered Formalin for possible histopathological examination. Select tissues (lungs, ovaries, testes, epididymides, Thyroid glands (including parathyroid glands), nasal cavity, trachea, bronchi, bone marrow (sternal and femoral sections and smear), prostate, scrotum, seminal vesicles, vagina, uterus, and other tissues with gross lesions from male and female rats of the control and high-dose groups were subjected to histopathologic examination. Testes and epididymides were fixed in Bouin’s fixative; sections were stained with periodic acid and Schiff’s (PAS) and counterstained with hematoxylin.

For animals dying during the study, a complete gross necropsy and histopathologic examination were conducted to determine the possible cause of death.

Statistics:

For body weight data, a repeated-measures analysis of variance (ANOVA) was conducted. An ANOVA with Bonferroni multiple comparisons was conducted on the thyroid hormone data. Fisher’s exact test was used on the reproductive data that are calculated on a group basis (e.g., gestational index). For reproductive data calculated on an individual basis (e.g., live birth index), micronuclei data, haematology and serum chemistry data, and organ weight data, the Wilcoxon rank sum test was used to increase the statistical sensitivity when comparing control group values to treated group values.

Clinical signs:

effects observed, non-treatment-related

Description (incidence and severity):

There were no treatment-related clinical findings. Areas of alopecia were sporadic and considered incidental.

Mortality:

no mortality observed

Description (incidence):

No deads reported

Body weight and weight changes:

effects observed, treatment-related

Description (incidence and severity):

Though not statistically significant, except for the final body weight values in female rats (day 93–95), there was a marginal decrease in mean body weights of the 2.0% group compared to the control group. The mild decrease in absolute body weights for this group of rats is due primarily to a depression in body weight gain during the first 2–3 wk of the study. Body weight values were normal for male and female rats of the 0.2 and 0.7% groups.

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):

For male and female rats, statistical significant differences in haematology parameters were few in number and not considered treatment related, due to (1) lack of consistency with time (7-wk compared to 14- wk values for male rats), (2) lack of dose response, or (3) the difference being of small magnitude (clinically insignificant). Decreases in total leukocyte count (14% lower than the control mean), haemoglobin (5%), and haematocrit (4%) were observed in the female rats of the 2.0% group.

Clinical biochemistry findings:

effects observed, treatment-related

Description (incidence and severity):

For male and female rats, statistical significant differences in serum chemistry parameters (except serum thyroid hormones) were few in number and not considered treatment related, due to (1) lack of consistency with time (7-wk compared to 14- wk values for male rats), (2) lack of dose response, or (3) the difference being of small magnitude (clinically insignificant). Increases in cholesterol (~30% higher than the control mean), total protein (5%), and albumin (9%) were observed in both the 0.7% and 2.0% groups of female rats.

Urinalysis findings:

not examined

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):

- Mean relative (to body weight) liver weights were mildly increased (6–10% above control values) in the male rats of the 2.0% group at both the 7-wk and 14-wk sacrifice periods. Mean absolute and relative epididymides weights were also increased in male rats of the 0.7% and 2.0% groups at 7 wk, but not at 14 wk. The increase in epididymides weights was not concentration dependent.
- For female rats, decreases in mean absolute organ weights included the brain (0.7% and 2.0%), ovaries (0.7% and 2.0%), and heart (2.0%). Mean relative brain (2.0%), kidney (2.0%), and liver (0.7%) weights were increased. Mean relative ovaries weights were decreased (0.7% and 2.0%). Note that the mean body weight was statistically significantly decreased in the 2.0% female rats compared to the control female rats.

Gross pathological findings:

no effects observed

Description (incidence and severity):

There were no clinically significant or exposure-related gross lesions in any of the study animals.

Neuropathological findings:

not examined

Histopathological findings: non-neoplastic:

no effects observed

Description (incidence and severity):

There were no lesions of clinical significance in any treated (2.0% group) or control group animals. Tissues from animals in the 0.7% or 0.2% groups were not examined microscopically, due to the lack of microscopic findings in rats of the control and 2.0% groups.

Histopathological findings: neoplastic:

no effects observed

Other effects:

effects observed, treatment-related

Description (incidence and severity):

Statistically significant and concentration-related increases in T4 and rT3 were observed in male and female rats after 7 or 14 wk of exposure. T3 levels were decreased in a concentration-related manner in these animals. For TSH, an increase was observed in exposed male rats after 7 wk of exposure, but there was no concentration relationship. After 14 wk of exposure, male and female rats of the 2.0% group had statistically significant increases in TSH levels compared to control rats. A marginal and statistically significant increase in TSH was observed in female rats of the 0.2% group, but this effect was not statistically significant in female rats at 0.7%.

Key result

Dose descriptor:

NOAEC

Effect level:

20 000 ppm

Based on:

test mat.

Sex:

male/female

Basis for effect level:

other: Highest dose tested

Dose descriptor:

NOEC

Effect level:

7 000 ppm

Based on:

test mat.

Sex:

male/female

Basis for effect level:

other: Serum TSH concentrations

Key result

Critical effects observed:

no

Analytical verification of chamber atmosphere

The test material was 99.7+%, which remained stable throughout the study period. Analytical chamber concentrations matched the target exposure concentrations of 0 (control), 0.2, 0.7, and 2.0%.

The nominal mean concentrations (standard deviation) were 2.08% (0.09), 0.72% (0.03), and 0.24% (0.015) for the target concentrations of 2.0%, 0.70%, and 0.20%. The ratio between analytical/nominal concentrations as calculated from the rotometer settings for the air and test substance were 0.97, 0.98 and 0.83 for the 2%, 0.7%, and 0.2% target concentrations, respectively.

Chamber temperature and relative humidity means for all exposure groups ranged from 73.5 to 74.2°F and 43.4 to 54.8%, respectively. The oxygen concentration of the 2.0% chamber was approximately 19%. The deviation in this study from the accepted 10–14 air changes per hour during animal exposure to 5 air changes per hour did not have an impact on exposure temperature, relative humidity, oxygen concentration, or substance concentration.

Conversion of NOAEC

2% test substance is corresponding to 20.000 ppm or 160254 mg/m3 (based on a molecular weight of 195.91)

Conclusions:

Exposure of 2.0% test substance vapor for approximately 14 weeks produced minimal general toxicity in Sprague-Dawley rats.

Executive summary:

A combined repeated dose/ reproductive screening study was performed. The purpose of this study was to determine and evaluate the potential of the substance to produce reproductive toxicity and to provide additional information on the effect of test substance exposure on the thyroid. Groups of 16 male and 16 female rats were exposed (6hr/day) to substance vapor at concentrations of 0 (control), 0.2, 0.7, and 2.0% using whole body inhalation chambers. Prior to mating, rats were exposed for 4 weeks (5 days/wk). Exposures were 7 days/wk during the periods of mating (2 wk), gestation (3 wk), and lactation (3 wk). First generation pups were not exposed to substance vapor. In parental animals, there were no clinical signs of toxicity except for a minimal decrease in mean body weight in female rats at 2.0%. At necropsy, gross findings, mean serum chemistry levels, mean haematology values mean bone marrow micronuclei scores (detailled in section 7.6.2), and mean organ weights were similar for all exposure groups, including the air control group. Statistically significant differences were considered incidental. There were no treatment-related histopathologic tissue findings, including the thyroid organ. Analysis of reproductive indices and parameters indicate that the substance is not a reproductive toxicant. Results of serum thyroid hormone levels (e.g., T3, T4, rT3, and TSH), indicated concentration-related increases in TSH, T4, and rT3. T3 levels were decreased. First generation pup survival and mean body weights were similar in all exposure groups, including the control. Exposure of 2.0% vapor for approximately 14 weeks produced minimal general toxicity in Sprague-Dawley rats. On the basis of serum TSH concentrations, the no-observable-effect-level (NOEL) is 0.7%. The NOAEC for systemic toxicity was set at 2% (or 160254 mg/m3 based on a molecular weight of 195.91).

.

Endpoint conclusion

Endpoint conclusion:

adverse effect observed

Dose descriptor:

NOAEC

160 254 mg/m³

Study duration:

subchronic

Species:

rat

System:

endocrine system

Organ:

thyroid gland

Repeated dose toxicity: inhalation - local effects

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed

Repeated dose toxicity: dermal - systemic effects

Endpoint conclusion

Endpoint conclusion:

no study available

Repeated dose toxicity: dermal - local effects

Endpoint conclusion

Endpoint conclusion:

no study available

Additional information

Two sub-chronic studies (Dodd 1999 is a follow up of Dodd 1997) are available on the substance. In addition, several sub-acute studies are available, which are included as supporting information.

Combined reproductive screening and sub-chronic repeated dose toxicity study (Key study)

A combined repeated dose/ reproductive screening study was performed (Dodd 1999). The purpose of this study was to determine and evaluate the potential of the substance to produce reproductive toxicity and to provide additional information on the effect of test substance exposure on the thyroid. Groups of 16 male and 16 female rats were exposed (6hr/day) to substance vapor at concentrations of 0 (control), 0.2, 0.7, and 2.0% using whole body inhalation chambers. Prior to mating, rats were exposed for 4 weeks (5 days/wk). Exposures were 7 days/wk during the periods of mating (2 wk), gestation (3 wk), and lactation (3 wk). First generation pups were not exposed to substance vapor. In parental animals, there were no clinical signs of toxicity except for a minimal decrease in mean body weight in female rats at 2.0%. At necropsy, gross findings, mean serum chemistry levels, mean haematology values mean bone marrow micronuclei scores (detailled in section 7.6.2), and mean organ weights were similar for all exposure groups, including the air control group. Statistically significant differences were considered incidental. There were no treatment-related histopathologic tissue findings, including the thyroid organ. Analysis of reproductive indices and parameters indicate that the substance is not a reproductive toxicant. Results of serum thyroid hormone levels (e.g., T3, T4, rT3, and TSH), indicated concentration-related increases in TSH, T4, and rT3. T3 levels were decreased. Exposure of 2.0% vapor for approximately 14 weeks produced minimal general toxicity in Sprague-Dawley rats. On the basis of serum TSH concentrations, the no-observable-effect-level (NOEL) is 0.7%. The NOAEC for systemic toxicity was set at 2% (or 160254 mg/m3 based on a molecular weight of 195.91).

Sub-chronic repeated dose toxicity study

A sub-chronic (90 day) repeated dose toxicity study incl. a 4 week interim sacrifice was performed (Dodd 1997). Fifteen male and 15 female rats were exposed 2 hr/day, 5 days/week, for up to 13 weeks (65 exposures over 90 days) to 0 (air control), 2, 4, or 8% test substance. Prior to initiation of the exposures, rats were acclimated to the nose-only chamber restraint system, breathing air only, 2 hr/day for 1 week. Five male and 5 female rats per group were necropsied after 4 weeks on study (20 exposures). Records for body weights (weekly) and signs of toxicity including mortality (twice daily) were maintained. The morning after the last scheduled exposure, euthanasia was per formed by CO, inhalation overdose followed by exsanguination. At the 4-or 13-week termination, gross pathology was performed and tissues were harvested for histopathologic examination. Wet tissue weights were determined. As a suspected target organ, the thyroid glands were carefully processed. At both the 4-and 13-week animal terminations, blood was drawn immediately prior to termination from the vena cava for haematology and clinical chemistry assays. Bone marrow cells were collected from the femur and smears were prepared from five rats/sex/group at both the 4- and 13-week animal terminations to investigate the mutagenic potential of the substance via induction of micronuclei in bone marrow polychromatic erythrocytes. Deaths observed in the 2 and 8% groups were attributed to accidents resulting from the restraint system employed. Hematologic alterations were minimal and considered insignificant. Increases in the frequency of micronucleated bone marrow polychromatic erythrocytes were observed in rats of all three groups. Serum chemistry alterations observed in rats of all exposure groups included decreases in T3 and increases in thyroglobulin, rT3, T4, and TSH. Relative organ weight increases (8% group) occurred in the brain, liver, and thyroid glands; decreases were observed in the thymus and testes. A decrease in relative thymus weights and an increase in relative thyroid weights were observed also in rats of the 2 and 4% groups. Histopathological findings included a mild inflammation in the nasal turbinates of rats exposed to 4 or 8%, mild atrophy and degeneration of the testes (4 and 8% groups), and a mild increase in thyroid follicular colloid content in rats of all exposure groups. Though NOAECs were observed for select target organs (e.g., nasal turbinates, testes), an overall LOAEC of 20000 ppm is derived based on effects observed in thyroid glands and bone marrow.

14 -day dose range finder

A 14-day dose range finder was performed (Dodd 1997). In this study 4 Groups of 5 male Fischer-344 rats each were exposed for 2 hours/day, 5 days/ week to 0 (air), 3, 6 or 12% test substance, resulting in a total of 10 exposures. Daily mean exposure concentrations were within 2% of target concentrations. The morning after the last exposure, euthanasia was performed by CO2 inhalation overdose followed by exsanguination. Toxicological evaluations included, the recording of signs of toxicity and body weights. At termination, gross pathology was performed on all animals. Blood was drawn for haematology and clinical chemistry assays. Blood was drawn for haematology and clinical chemistry assays that included red blood cell count, haemoglobin. mean corpuscular volume, mean corpuscular haemoglobin, mean corpuscular haemoglobin concentration. haematocrit, total and differential leukocyte count. total protein, albumin, globulin, alanine aminotransaminase (ALT), aspartate aminotransaminase (AST), thyroxine (T4), and thyroxine binding globulin. In addition, wet tissue weights (lungs, heart, liver, kidney, testes, and thyroid/parathyroid), serum triiodothyronine (T3), thyroglobulin, thyroid-stimulating hormone (TSH), reverse T3 (rT3), free T4 index, and platelet count were determined. The thyroid and parathyroid glands were harvested for histopathologic evaluation. Evaluation of organ weights, gross lesions and histopathology did not reveal any abnormalities. Mild effects on clinical signs and weight gain and decreased WBC were observed in the 6 and 12% groups. decreased albumin was observed in the 12% group and Serum thyroglobulin and reverse T3 values were significantly increased in all exposure groups. This data is used to determine the dose range of the sub-chronic study.

Sub-acute study (OECD 412)

The inhalation toxicity of the substance was studied in a sub-acute (4-week) study in Wistar rats, according to OECD guideline 412 and GLP principles (TNO 2007). Groups of 5 male and 5 female rats were exposed nose-only to target concentrations of 0 (clean air control) 10,000 ppm, 20,000 ppm, or 40,000 ppm for 6 hours a day, 5 days a week during a 4-week period, with a total number of 20 exposure days. To examine the toxicity of the test material, data on clinical observations, body weight gain, food consumption, food conversion efficiency, haematology and clinical chemistry were used. In addition, animals were examined grossly at necropsy, organs were weighed, and a selection of organs and tissues (Including the complete respiratory tract and nasal passages) was examined microscopically, and the bone marrow of all male animals was used for analysis of chromosomal aberrations. Finally, additional groups of male animals exposed to the control condition, 20,000 or 40,000 ppm were subjected to liver perfusion at necropsy to harvest cells for an Unscheduled DNA Synthesis (UDS). These genotoxicity endpoints are summarized separately in section 7.6.2. Daily observations of the animals before each exposure or halfway the exposure period did not reveal any abnormalities. However, on several days during the first two weeks of exposure, after each day's exposure, clinical signs such as piloerection, coprophagy, and reduced food intake were observed and animals felt cold upon touch. These signs were seen m all groups, controls included. Seven days after exposure start, reduced body weight was seen in male animals exposed to 20,000 or 40,000 ppm. Thereafter weight gain was observed, however, body weights were still significantly lower than in controls 14 days after exposure start (20,000 ppm group), and 14, 21 and 27 days after exposure start (40,000 ppm group). Some body weight decrease was also seen during the first week in all female groups, but this was also seen in controls. In the first exposure week, food consumption was decreased in males and females exposed to 20,000 or 40,000 ppm; it recovered thereafter in female animals whereas food intake was still lower in these male groups when compared to controls. Food conversion efficiency was negative in males exposed to 20,000 or 40,000 ppm and in all female groups (including controls) during the first week. Statistically significant changes in red blood cell parameters consisted of a decrease m thrombocytes in females exposed to 20,000 or 40,000 ppm, a decrease in mean corpuscular haemoglobin concentration (MCHC) in females exposed to 40,000 ppm, and a decrease in haemoglobin concentration in all exposed female groups. A clear concentration-related decrease in haemoglobin concentration in these animals was, however, not observed. No such changes were observed in males. Decreased white blood cell numbers were observed in animals exposed to 40,000 ppm; a statistically significant degree was almost reached in male rats. Statistically significant changes in clinical chemistry parameters included a decrease in cholesterol in male and female animals exposed to 40,000 ppm, an increase in glucose, alkaline phosphatase (ALP), and total bilirubin in female animals exposed to 20,000 or 40,000 ppm, and an Increase in alanine aminotransferase (ALAT), urea and triglycerides in females exposed to 40,000 ppm. Such increases could also be observed m males exposed to 40,000 ppm, but a statistically significant level was not reached. An increase in albumin/globulin ratio was seen in males and females exposed to 40,000 ppm; in males it reached statistical significance, in females it was accompanied by significant decreases in albumin and total protein. Absolute and relative spleen weight was decreased In males exposed to 20,000 or 40,000 ppm, and relative spleen weight in females exposed to 40,000 ppm. Absolute liver weight was increased in females exposed to 20,000 or 40,000 ppm; relative liver weight was increased in males and females exposed to 40,000 ppm. Macroscopic examination at necropsy revealed a smaller thymus in all males and two females exposed to 40,000 ppm. Histopathological treatment-related changes were found in the adrenals and the spleen of males and females exposed to 40,000 ppm, and in the testes of males exposed to 40,000 ppm. These changes consisted of increased vacuolation in the zona fasciculate of the adrenals, which was slightly more prominent in males than in females, very slight to moderate lymphoid depletion of the periarterial lymphocytic sheath (PALS) of the spleen, and a few multinucleated giant cells in the testes, indicative of germ cell degeneration. The changes in the thymus of all males and two females exposed to 40,000 ppm (collected at necropsy because of gross lesions) revealed a moderately to severely decreased size of the medulla, in some cases accompanied by an increased number of necrotic lymphocytes. No other exposure-related findings were observed. In the CA assay the substance was found to be cytotoxic to the bone marrow in a concentration-related way. In conclusion, because of the changes induced in lymphoid organs, the should be considered an immunotoxic compound. A LOAEL of 10.000 ppm is derived based on the cytotoxicity in bone marrow in all dose groups.

Sub-acute study (OECD 412) in mice with additional sub-groups for thyroid hormone and serum iodine analysis

A non-GLP sub-acute repeated dose exposure study was performed in compliance with OECD guideline 412 (Lovelace 2017). The purpose of this study was to test tolerance and thyroid response to whole body exposure of the test substance in B6C3F1 male mice. Groups of randomized mice (N=42) were exposed to filtered air, or test substance gas at a target concentration of 2,500 ppm (0.25%), 5,000 ppm (0.5%) or 10,000 ppm (1.0%) for up to 28 consecutive days (6 hours/day) starting at study day (SD) 1. Subgroups of mice from each exposure group were euthanized for evaluation at SD 14 (after 13 days of exposure, referred to as “TH1 mice” n=18/group), SD 28 (after 27 days of exposure, referred to as “TH2 mice” n=12/group), and SD 29 (after 28 days of exposure, referred to as “main study mice” n=6/group). The remaining 6 mice in each group (referred to as “TH2R mice”) were euthanized at SD 44 after recovery for 17 days following the 27 days of exposure. Clinical signs and body weights were evaluated throughout the study. Body temperature was measured for main study mice pre- and post- exposure on SD 1, 3, 14 and 28. At SD 14, 28 and 44, serum thyroid hormones (T3, T4 and TSH) and iodide (SD 14 only) concentration were measured; thyroid and liver weights were also collected. At SD 29, clinical pathology and complete gross pathology, as well as histopathology and weight of thyroid, were evaluated. The exposure atmosphere was monitored using a hydrocarbon analyzer with Flame Ionization Detector. The average measured CF3I concentration during the 28 day exposure for the low, mid and high dose group was 2,531 ±30 ppm, 5,062±72 ppm, and 10,011±94 ppm, respectively. No adverse clinical signs were observed in any animals throughout the study. There were no exposure associated changes in body weights and body temperature in mice exposed to CF3I, compared to the air control group. Circadian rhythm variation of body temperature was observed in all mice including air control animals. An exposure concentration-related increase in absolute and relative liver weight was identified in all exposure dose groups after 13 days of exposure, but the increase was limited to relative liver weight in the high dose group after 27 days of exposure. The liver weight changes were completely resolved by SD 44 (i.e. after a 17-day recovery period post 28 days of exposure). Since no adverse clinical signs were observed on these animals and clinical pathology data did not demonstrate changes in parameters associated with liver toxicity, the transient, recoverable liver weight changes are considered to be non-adverse, although histopathology of liver was not evaluated. As to the thyroid parameters, serum iodide levels were similar to the control mice in the exposed mice after 13 days of exposure. Serum T4 levels in the mid and high dose group mice at SD 14 were found to be higher than air control mice but were similar to control levels when measured at SD 28. Serum T3 levels were significantly greater than air controls at the high dose but were similar to controls at the mid and low dose at SD 28. In addition, serum TSH levels were slightly, but statistically significantly lower in the mid dose group compared to air control group at SD 14 but not different from controls at SD28. All three thyroid hormone levels were similar among all dose groups at SD 44 after a 17-day recovery period, suggesting the effects on thyroid hormones were transient and recoverable. In addition, there was equivocal evidence that thyroid gland weights were increased in the high dose group as the difference achieved statistical significance at SD 28 (after 27 days of exposure), but not at SD 29 (after 28 days of exposure). Thyroid gland weights were comparable among all dose groups at SD 44 after a 17-day recovery period following 28 days of exposure. No gross lesions were found when complete necropsy was performed on main study mice after 28 days of exposure. Microscopic examination on thyroid glands from main study mice did not reveal any exposure associated changes. In conclusion, exposure to 2,500 ppm (0.25%), 5,000 ppm (0.5%), and 10,000 ppm (1.0%) for consecutive 28 days (6 hours/day) produced no overt general toxicity and only transient, recoverable effects on thyroid weight and hormones and liver weight. On the basis of the effect of exposure on the thyroid including evaluation of thyroid histopathology, the NOAEL for this study is 10,000 ppm (1.0%).

Sub-acute study according to Japanase guidelines

In a sub-acute (28 day) whole body inhalation study, performed according to Japanese guidelines, Crj: CD(SD) rats (6/dose/sex) were exposed to 0.27, 1.01 and 5.28 mg/L (analytical concentration, corresponding to nominal concentrations of 0.2, 1.0 and 5.0 mg/L test substance (Mitsubishi 2001). In clinical observation, body weight shift, food consumption, haematological examination and blood biochemical examination, no change related to the test substance was observed. In organ weights, though the weight changes of spleen, brain and liver were observed in animals of 5.0 mg/L group, any histopathological changes due to test material were not observed. It was concluded that test material did not cause these changes. Based on these results, the substance did not induce any toxicological changes to rat under these experimental conditions. Accordingly the NOAEL is exceeding 5.28 mg/L. 

Justification for classification or non-classification

Based on the results of the sub-chronic repeated dose toxicity studies in rats, classification and labelling for toxicity upon repeated or prolonged exposure is not warranted in accordance with EU Classification, Labelling and Packaging of Substances and Mixtures (CLP) Regulation (EC) No. 1272/2008 and its updates.