Hexafluoropropene

Endpoint summary

Key value for chemical safety assessment

Effects on fertility

Description of key information

- Inhalation, rat, OECD 443, (P) Systemic NOAEC: 50 ppm; Reproductive toxicity NOAEC: => 600 ppm, (F1) Systemic NOAEC: 50 ppm; Sexual development NOAEC: => 200 ppm, Triskelion 2018

Effect on fertility: via oral route

Endpoint conclusion:

no study available

Effect on fertility: via inhalation route

Endpoint conclusion:

no adverse effect observed

Study duration:

subchronic

Species:

rat

Quality of whole database:

GLP compliant OECD 443 study

Effect on fertility: via dermal route

Endpoint conclusion:

no study available

Additional information

Extended One Generation Reproducitve Toxicity Test - Rat

The objective of the GLP-compliant study, performed according to OECD 443, was to provide data on the possible effects of the test substance on reproductive performance of Wistar rats and the development of pups consequent to daily inhalation exposure to various concentrations of the test substance (intended target concentrations: 0 ppm, 50 ppm, 300 ppm and 900 ppm) to male and female rats during a premating period of 10 weeks and during mating (max. 2 weeks), gestation and lactation until postnatal day (PN) 21. At weaning on postnatal day 21 (PN 21), pups were distributed to Cohorts 1A and 1B and were exposed to the test substance at lower concentrations as their parents during their growth into adulthood (intended target concentrations: 0 ppm, 50 ppm, 300 ppm and 600 ppm).

The overall average actual concentration (± standard deviation) of the test substance in the low-concentration test atmosphere (group 2; 50 ppm target concentration), as determined by total carbon analysis, was 49.3 (± 2.1) ppm. Exposure to the mid-concentration (group 3) was started at a target concentration of 300 ppm, which was reduced to 100 ppm as of 3 June 2018; the average actual concentrations were 300.3 (± 7.9) and 100.3 (± 1.2) ppm, respectively. Average concentrations in the high-concentration test atmosphere (group 4) were 900.3 (± 12.9), 601.0 (± 19.5) and 199.8 (± 2.4) ppm, in the periods 5-9 February, 10 February — 2 June, and 3 June — 29 August 2018 when target concentrations were 900, 600 and 200 ppm, respectively.

Three P0-females of the high-concentration group (900 ppm) and several animals of Cohorts 1A and 1B of the high-concentration groups (600 ppm) died during the first days of exposure. Accordingly, the target concentration of the high-concentration group was lowered during exposure of the P0-generation (from 900 ppm to 600 ppm) and the target concentrations of the mid- and high-concentration groups of the F1-generation were lowered before the start (day 0) of Cohort 1A and Cohort 1B (from 300 ppm to 100 ppm and from 600 ppm to 200 ppm, respectively). Animals that died were replaced by surplus animals and after lowering the target concentrations, no other animals died during the study.

Clinical observations revealed piloerection, hunched posture, respiration dyspnoea and/or muscle weakness in several animals of the high-concentration group during the first exposure days of the P0-generation and in the first exposure weeks of Cohort 1A and 1B. These signs were mainly observed in the high-concentration group (900 ppm) of the P0-generation and in the high-concentration group (600 ppm) of Cohorts 1A and 1B, before lowering the target concentrations and disappeared thereafter. No other treatment-related (detailed) clinical signs were observed during the study.

In the P0-generation, body weights of the male animals of the mid- and high-concentration groups and in females of the high-concentration group were statistically significantly lower than the corresponding control animals (maximally, during premating: P0-males 17%, P0-females 10%; P0-females during gestation 9%, P0-females during lactation 7%).

In Cohorts 1A and 1B of the F1-generation, body weights were statistically significantly lower in the males and females of the mid- and high-concentration groups (in Cohort 1A maximally 30% in males and 28% in females, respectively, and in Cohort 1B maximally 35% in males and 30% in females, respectively).

In the P0-generation, food consumption of the male and female animals of the high-concentration group was statistically significantly lower than of the control animals (maximally 44% in males and 39% in females). In male and females animals of the mid-concentration groups food consumption was statistically significantly lower compared to control animals during the first week(s) of the study.

In the F1-generation, food consumption of the male and females animals of the high-concentration group was statistically significantly lower than of controls (in Cohort 1A maximally 25% in males and 18% in females, respectively, and in Cohort 1B maximally 22% in males and 16% in females, respectively).

The observed effects on body weights were considered, at least in part, as transient and/or as related to the lower food consumption. The observed effects on body weights in the high-concentration groups of the P0- (600 ppm) and F1- (200 ppm) generations, however, were considered to be related to treatment.

Except for the increase in urea in male animals of the high-concentration group of the P0-generation (68% higher than controls) and in male animals of the high-concentration group of Cohort 1A of the F1-generation (18% higher than controls), no treatment-related effects were observed on haematology and clinical chemistry parameters.

Urinalysis revealed a higher urinary volume in males of the low-, mid-and high-concentration groups of the P0-generation and in females of the high-concentration group of Cohort 1A of the F1-generation.

In the P0-generation, the increased relative kidney weight in male and female animals of the mid-and high-concentration groups (300-600 ppm: ~91-130% increase in males and ~44­65% increase in females compared to controls, respectively) were considered to be adverse and related to treatment. Additionally, in P0-generation animals, terminal body weight was decreased in males of the mid- and high-concentration groups (300 and 600 ppm; maximally 17% below controls). Furthermore, the increased relative lung weight (~19-30% in males and ~14% in females), heart weight (~20-39% in males and ~22% in females), liver weight (~15-18% in males and ~14% in females), and spleen weight (~10-21% in males and ~13% in females) as observed in male animals of the mid- (300 ppm) and high-concentration (600 ppm) groups and in female animals of the high-concentration group (600 ppm) were considered to be related to treatment.

In Cohort 1A of the F1-generation, the increased relative kidney weight in male and female animals of the mid-and high-concentration groups (100-200 ppm: ~17-58% increase in males and ~20-51% increase in females compared to controls, respectively) were considered to be adverse and related to treatment. Additionally, in Cohort 1A animals, terminal body weight was decreased in high-concentration (200 ppm) male and female animals (decrease of 13% and 12%, respectively). Furthermore, the increased relative lung weight (~7-13% in males and ~6-10% in females) and relative liver weight (~9-13% in males and ~7-11% in females) as observed in males and females of the mid- (100 ppm) and high concentration (200 ppm) groups and the increased relative heart weight observed in high-concentration (200 ppm) males and mid- (100 ppm) and high-concentration (200 ppm) females (~13% and 15%, respectively) were considered to be related to treatment.

No effects were observed on the weight of the reproductive organs of Cohort 1B F1-generation animals.

At necropsy, macroscopic examination revealed pale discoloration, enlargement and/or a pitted surface of the kidneys in mid- and high-concentration animals of the P0-generation (300 ppm and 600 ppm) and in animals of Cohort 1A of the F1-generation (100 ppm and 200 ppm).

Microscopic examination of the sampled organs and tissues in the P0-generation animals revealed treatment-related histopathological changes in the heart (minimal to moderate ventricular muscle degeneration) in high-concentration females (600 ppm) and kidneys (minimal to moderate tubular dilatation, mononuclear inflammation, proteinaceous casts and/or basophilic tubules) in mid- and high-concentration animals (300 and 600 ppm, respectively).

Microscopic examination of the sampled organs and tissues of the animals of Cohort 1A of the F1-generation revealed treatment related histopathological changes in the kidneys (minimal to moderate tubular dilatation, mononuclear inflammation, proteinaceous casts and/or basophilic tubules) in mid- and high-concentration animals (100 and 200 ppm, respectively).

Other organs and tissues did not reveal treatment related histopathological changes.

No treatment-related effects were observed on the fertility and reproductive

performance of male and female animals of the P0-generation.

No treatment-related effects were observed on estrous cycle related parameters in female animals of the P0-generation and in animals of Cohort 1A of the F1-generation.

No treatment-related effects were observed on epididymal and testicular sperm parameters in male animals of the P0-generation and in animals of Cohort 1A of the F1-generation.

No effects were observed on TSH and T4 analysis in animals of the P0-generation and in adult F1-generation animals of Cohort 1A.

No treatment-related effects were observed on number of live pups, number of implantation sites, implantation loss, stillborn pups, dead, missing and/or cannibalized pups, litter loss, pup viability indices and sex ratio.

No treatment-related effects were observed on clinical signs of pups nor on macroscopic observations at sacrifice and of dead pups in F1-generation pups.

Overall, in the mid- (300 ppm) and high-concentration group (600 ppm), the body weight of F1-generation pups during the lactation period was lower than of the corresponding control pups (~10-15% on postnatal day 21). This finding was considered to be related to treatment.

No direct effects were observed on anogenital distance on PN4 in F1-generation pups or on nipple retention in male F1-generation pups.

Preputial separation (control: 42.4 days, high dose 46.4 days) was delayed in male pups of the high-concentration group (200 ppm). However, these differences were not considered as delayed sexual development but as a consequence of delayed general development (lower pup weights).

At sacrifice of the selected pups at post-natal day 21, the terminal body weights of the pups of the mid-concentration (300 ppm) and high-concentration (600 ppm) groups calculated per litter were lower than of the control group. No direct effects were observed on organ weights of F1-generation pups sacrificed on post-natal day 21.

No treatment-related effects were observed on the development of the ovarian follicles from primordial small follicles into corpora lutea in Cohort 1A animals of the F1-generation.

No treatment-related effects were observed on splenic lymphocyte subpopulation analysis in Cohort 1A animals of the F1-generation.

Based on the decreased body weights (change), decreased food consumption, increased urinary volume, effects on organ weights, macroscopic observations in the kidneys and microscopic effects in several organs (kidneys, heart and thymus) the No Observed Adverse Effect Concentration (NOAEC) for parental toxicity was placed at the low-concentration (50 ppm) after exposure via inhalation for 6 hours/day, 7 days/week for up to 12 weeks for males (pre-mating through mating) and for 12 weeks for females (pre-mating through mating) and then through post-natal day (PND) 21.

There were no treatment-related adverse effects of exposure to the test substance on any of the multiple endpoints assessed with regards to fertility and reproductive performance in the parental generation (P0-animals). In addition, there also were no treatment-related adverse effects on the various reproductive systems endpoints assessed in the male and female F1-animals (Cohorts 1A and 1B). Based on the absence of any treatment-related adverse effects on any fertility and reproductive performance parameters in this OECD 443-compliant study, the study NOAEC for fertility and reproductive performance was placed at the high-concentration (600 ppm).

Based on the increased relative organ weights (kidney, lung, liver, heart) and the histopathological changes in the kidney after macroscopic and microscopic examination in mid-(100 ppm) and high-concentration (200 ppm) animals of Cohort 1A, the No Observed Adverse Effect Concentration (NOAEC) was based on these general toxicity observations and placed at the low-concentration (50 ppm). There were no treatment-related effects on sexual development at any exposure concentration in the F1-generation animals.

Effects on developmental toxicity

Description of key information

- Inhalation, rat, OECD 414, (P) Systemic NOAEC: 300 ppm, (F1) Developmental NOAEC: 300 ppm, Triskelion 2018

- Inhalation, rabbit, OECD 414, (P) Systemic NOAEC: =>300 ppm, (F1) Developmental NOAEC: =>300 ppm, Triskelion 2019

Effect on developmental toxicity: via oral route

Endpoint conclusion:

no study available

Effect on developmental toxicity: via inhalation route

Endpoint conclusion:

no adverse effect observed

Study duration:

subacute

Species:

rat

Effect on developmental toxicity: via dermal route

Endpoint conclusion:

no study available

Additional information

Developmental toxicity study - Rat

The objective of this GLP compliant OECD 414 study was to provide data on the possible effects of the test substance on pregnant female Wistar rats, and on the development of the embryo and fetus. The test material was administered by inhalation (whole body exposure) to groups of 24 mated females from gestation day (GD) 6 up to and including GD 20 at concentrations of 50, 300 and 900 ppm. Verification of the target concentrations were performed throughout exposure. Maternal rats were observed for manifestation of clinical signs before, during and after exposure. Body weights of the parental female animals was recorded on gestation days (GD) 0, 6, 9, 12, 15, 18 and 21. Food consumed for each mated female was measured. The females were killed by exsanguination after isofluorane anaesthesia on GD 21 and examined for gross abnormalities, organ weights and uterine content. Fetuses were examined for external alterations and sacrificed by hypothermia. Subsequently, approximately half of the fetuses of each litter were examined for soft tissue anomalies and the other half were examined for skeletal abnormalities.

The overall average actual concentrations (± standard deviation) of test substance in the test atmospheres, as determined by total carbon analysis, were 50 (± 0.7), 302 (± 3) and 901 (± 16) ppm for the low-, mid- and high-concentration, respectively. These concentrations were close to the respective target concentrations of 50, 300 and 900 ppm. At gestation day 21 caesarean section was performed, dams were examined macroscopically and reproductive organs, liver and kidney were weighed. Fetuses and placentas were weighed and fetuses were examined externally. The exposure to the test material was well tolerated at low and mid concentration and did not induce treatment-related changes in general condition. Females in the high concentration group frequently showed piloerection. Statistically significantly reduced body weight and body weight gain was observed in the females of the mid and high concentration group. Mean body weight at the end of gestation in the mid concentration group was less than 5% lower as compared to the control group. This was considered a slightly lower body weight and not considered to be adverse. Mean body weight at the end of gestation in the high concentration group was 15% lower as compared to the control group. This was considered an adverse and treatment-related effect. Concentration related lowered mean food consumption was observed in the mid and high concentration groups for intervals 6-9 days and later. Uterus weight for both full and empty uterus was lower in the high concentration group, which could be related to maternal toxicity. Organ weights revealed an increase in weight of the kidneys, visible already at the 50 ppm concentration. Macroscopic pathologic observation of the kidneys showed enlargement and discoloration in the 900 ppm group. In combination with the effects on kidney weight this was considered a treatment-related and adverse effect.

The slightly lower body weight and feed intake observed in the 300 ppm group were considered related to treatment, but in view of the severity considered not adverse. The observed effects on body weight, feed intake and kidney weight (supported by macroscopic observations in the 900 ppm group) were considered related to treatment and adverse. Therefore, the No Observed Adverse Effect Concentration(NOAEC)for maternal toxicity was placed at 300 ppm.

Mean fetus weight was statistically significantly lower in the mid and high concentration groups. A concentration-dependent effect was observed with less than 5% lower fetus weight in the 300 ppm group and more than 15% lower fetus weight in the 900 ppm group as compared to the control group. Furthermore, a number of skeletal observations indicative of retardation in ossification were noted. With some exceptions, statistical significance was mostly confined to the high-concentration group. The retardation in ossification is considered to be related to the lower fetus weight observed.

The slight effect on fetus weight and ossification in the 300 ppm group was considered related to treatment, but in view of the severity were not considered to be biologically relevant and, therefore, not adverse. The effect on fetus weight, accompanied by a retardation in ossification observed in the 900 ppm group was considered adverse. Therefore NOAEC for developmental toxicity was placed at 300 ppm.

 

Developmental toxicity study - Rabbit

In this GLP compliant OECD 414 study, the objective was to provide data on the possible effects of the test substance on pregnant female New Zealand White rabbits and the development of the embryo and fetus consequent to daily inhalation exposure to the test substance from gestation day (GD) 6 up to and including gestation day 28.

Animals were exposed to test atmospheres in whole body exposure chambers for 6 hours per day at concentrations of 0, 10, 50 and 300 ppm test substance.

Each group comprised of 22 mated females and in-life parameters included mortality and morbidity, body weight and food consumption. At gestation day 29 caesarean section was performed on the animals in all groups and necropsy parameters included examination of the dams for gross anatomical changes, uterus and ovary weight. The number and distribution of implantation sites, live and dead fetuses and resorptions were recorded. In addition placentas and live fetuses were weighed individually.

Fetuses from all groups were examined for external and visceral malformations. After visceral examination, the fetal bodies were processed and stained with Alzarin Red S. for skeletal examination. Heads of half of the fetuses in each litter were fixed in Bouin’s fixative for visceral head examination.

Daily exposure to 0, 10, 50 and 300 ppm test substance from gestation day 6 up to and including day 28 in New Zealand White rabbits resulted in:

- One animal in the 10 ppm group died spontaneously. No other mortality or treatment-related morbidity were observed.

- Statistically significant, but not biologically adverse lower body weight was observed in the 300 ppm group at days 12 and 15 compared to the control group

- Statistically significant, but not biologically adverse lower body weight gain was observed as compared to the control group from GD 6-9 and GD 9-12 in the high concentration group (300 ppm).

- Food intake was decreased in the high concentration group from GD 6-9, GD 9-12 and GD 12-15, whereas in the low and mid concentration groups no effects were observed on body weight or food consumption.

- No treatment-related maternal macroscopic deviations were observed.

- No effects on the mean number of corpora lutea, the number and distribution of implantation sites, live or dead fetuses, and early and late resorptions were observed.

- No effects on the mean fetus weight in both male and female fetuses were observed

- Fetal external and visceral examinations showed no treatment-related effects.

- A slight retardation in ossification was observed in fetuses in the 300 ppm group.This was considered a treatment-related effect, but not adverse.

 

CONCLUSIONS

There were no treatment-related adverse effects in the pregnant females at any concentration. Therefore the No Observed Adverse Effect Concentration for maternal toxicity was placed at 300 ppm test substance.

Delayed ossification (a variation) was observed at 50 ppm, but there were no treatment-related adverse effects in the fetuses at any concentration. Therefore, the No Observed Adverse Effect Concentration for developmental toxicity was placed at 300 ppm.

Justification for classification or non-classification

Based on the available information, classification for reproductive toxicity is not warranted in accordance with EU Classification, Labelling and Packaging of Substances and Mixtures (CLP) Regulation No. (EC) 1272/2008.