cis-2-methyl-4-propyl-1,3-oxathiane

Toxicological Summary

Workers - Hazard via inhalation route

Systemic effects

Long term exposure

Hazard assessment conclusion:

DNEL (Derived No Effect Level)

Value:

3.53 mg/m³

Most sensitive endpoint:

effect on fertility

Route of original study:

Oral

DNEL related information

DNEL derivation method:

ECHA REACH Guidance

Overall assessment factor (AF):

75

Dose descriptor starting point:

NOAEL

Value:

300 mg/kg bw/day

Modified dose descriptor starting point:

NOAEC

Value:

264.5 mg/m³

Explanation for the modification of the dose descriptor starting point:

Regarding absorption, in the absence of reliable data for both the starting route (oral) and the end route (inhalation), worst-case assumptions were made. It was assumed that a limited absorption occurs by the oral route, leading to a low (conservative) internal NOAEL. To secure a conservative external NOAEL, a maximum absorption should be assumed for the inhalation route (i.e.; 100%) leading to a low external NOAEL. Thus, in the case of oral-to-inhalation extrapolation, it is proposed to include a default factor of 2, i.e. the absorption percentage by oral route is half that of the inhalation absorption as suggested in ECHA Guidance on Information Requirements and Chemical Safety Assessment, Chapter R.8 (2012).

To convert the oral NOAEL into inhalatory NOAEC, a rat default respiratory volume was used corresponding to the daily duration of human exposure (sRVrat: 0.38 m3/kg bw/8 h). For workers a correction was added for the difference between respiratory rates under standard conditions (sRVhuman: 6.7 m3 for an 8 h exposure period) and under conditions of light activity (wRV: 10 m3 for an 8 h exposure period).

The corrected dose descriptor for inhalation is determined using the following equation:

Corrected Inhalatory NOAEC = [NOAEL] X (1/SRVrat x ABS(oral-rat)/ABS(inh-human) x sRVhuman/wRV

= [300 mg/kg bw/day] X  [1/0.38 m3/kg bw/day] X [1/2] X [6.7 m3/10m3].

Thus, the corrected dose descriptor for inhalation is 264.5 mg/m3 for workers.

AF for dose response relationship:

1

Justification:

The dose-descriptor is a NOAEL. Table R.8-6 ECHA REACH Guidance.

AF for differences in duration of exposure:

6

Justification:

Default factor for a sub-acute study. Table R.8-5 ECHA REACH Guidance.

AF for interspecies differences (allometric scaling):

1

Justification:

Table R.8-4 ECHA REACH Guidance. Assessment factor not to be used for inhalation route since the differences in the metabolic rate/bw has already been taken into account in the corrected dose descriptor.

AF for other interspecies differences:

2.5

Justification:

Default factor for other interspecies differences. Table R.8-6 ECHA REACH Guidance

AF for intraspecies differences:

5

Justification:

Default factor for worker. Table R.8-6 ECHA REACH Guidance.

AF for the quality of the whole database:

1

Justification:

Default factor for good/standard quality of the database has taken into account completeness of the standard information requirements for the tonnage band.

AF for remaining uncertainties:

1

Justification:

No remaining uncertainties

Acute/short term exposure

Hazard assessment conclusion:

DNEL (Derived No Effect Level)

Value:

10.59 mg/m³

Most sensitive endpoint:

repeated dose toxicity

Route of original study:

Oral

DNEL related information

DNEL derivation method:

ECHA REACH Guidance

Overall assessment factor (AF):

75

DNEL extrapolated from long term DNEL

Dose descriptor starting point:

NOAEL

Value:

300 mg/kg bw/day

Modified dose descriptor starting point:

NOAEC

Value:

793.5 mg/m³

Explanation for the modification of the dose descriptor starting point:

Regarding absorption, in the absence of reliable data for both the starting route (oral) and the end route (inhalation), worst-case assumptions were made. It was assumed that a limited absorption occurs by the oral route, leading to a low (conservative) internal NOAEL. To secure a conservative external NOAEL, a maximum absorption should be assumed for the inhalation route (i.e.; 100%) leading to a low external NOAEL. Thus, in the case of oral-to-inhalation extrapolation, it is proposed to include a default factor of 2, i.e. the absorption percentage by oral route is half that of the inhalation absorption as suggested in ECHA Guidance on Information Requirements and Chemical Safety Assessment, Chapter R.8 (2012).

To convert the oral NOAEL into inhalatory NOAEC, a rat default respiratory volume was used corresponding to the daily duration of human exposure (sRVrat: 0.38 m3/kg bw/8 h). For workers a correction was added for the difference between respiratory rates under standard conditions (sRVhuman: 6.7 m3 for an 8 h exposure period) and under conditions of light activity (wRV: 10 m3 for an 8 h exposure period).

The corrected dose descriptor for inhalation is determined using the following equation:

Corrected Inhalatory NOAEC = [NOAEL] X (1/SRVrat x ABS(oral-rat)/ABS(inh-human) x sRVhuman/wRV = [300 mg/kg bw/day] X  [1/0.38 m3/kg bw/day] X [1/2] X [6.7 m3/10m3].

The extrapolation of long to short term risk was set based on a reference period of 15 minutes at 1-5 times the value (default 3) of the long-term DNEL i.e. Corrected Inhalatory NOAEC x 3.

Thus, the corrected dose descriptor for inhalation is 793.5 mg/m3 for workers.

AF for dose response relationship:

1

Justification:

The dose-descriptor is a NOAEL. Table R.8-6 ECHA REACH Guidance.

AF for interspecies differences (allometric scaling):

1

Justification:

Table R.8-4 ECHA REACH Guidance. Assessment factor not to be used for inhalation route since the differences in the metabolic rate/bw has already been taken into account in the corrected dose descriptor

AF for other interspecies differences:

2.5

Justification:

Default factor for other interspecies differences. Table R.8-6 ECHA REACH Guidance

AF for intraspecies differences:

5

Justification:

Default factor for worker. Table R.8-6 ECHA REACH Guidance.

AF for the quality of the whole database:

1

Justification:

Default factor for good/standard quality of the database has taken into account completeness of the standard information requirements for the tonnage band.

AF for remaining uncertainties:

10

Justification:

Default factor for a sub-acute study. Table R.8-5 ECHA REACH Guidance.

Local effects

Long term exposure

Hazard assessment conclusion:

no hazard identified

Acute/short term exposure

Hazard assessment conclusion:

no hazard identified

DNEL related information

Workers - Hazard via dermal route

Systemic effects

Long term exposure

Hazard assessment conclusion:

DNEL (Derived No Effect Level)

Value:

1 mg/kg bw/day

Most sensitive endpoint:

effect on fertility

Route of original study:

Oral

DNEL related information

DNEL derivation method:

ECHA REACH Guidance

Overall assessment factor (AF):

300

Dose descriptor starting point:

NOAEL

Value:

300 mg/kg bw/day

Value:

300 mg/kg bw/day

Explanation for the modification of the dose descriptor starting point:

For systemic hazard assessment via the dermal route of exposure, route-to-route extrapolation from the oral NOAEL value was considered appropriate. As no data on dermal penetration are available, dermal absorption is considered to be the same as oral absorption (i.e. 100 %). Therefore the oral NOAEL is considered the same as the dermal NOAEL (ECHA Guidance on Information Requirements and Chemical Safety Assessment, Chapter R.8 (2012)).

AF for dose response relationship:

1

Justification:

The dose-descriptor is a NOAEL. Table R.8-6 ECHA REACH Guidance.

AF for differences in duration of exposure:

6

Justification:

Default factor for a sub-acute study. Table R.8-5 ECHA REACH Guidance.

AF for interspecies differences (allometric scaling):

4

Justification:

Default allometric scaling factor for rats. Table R.8-4 ECHA REACH Guidance.

AF for other interspecies differences:

2.5

Justification:

Default factor for other interspecies differences. Table R.8-6 ECHA REACH Guidance.

AF for intraspecies differences:

5

Justification:

Default factor for worker. Table R.8-6 ECHA REACH Guidance.

AF for the quality of the whole database:

1

Justification:

Default factor for good/standard quality of the database taken into account completeness of the standard information requirements for the tonnage band.

AF for remaining uncertainties:

1

Justification:

No remaining uncertainities.

Acute/short term exposure

Hazard assessment conclusion:

no hazard identified

DNEL related information

Local effects

Long term exposure

Hazard assessment conclusion:

no hazard identified

Acute/short term exposure

Hazard assessment conclusion:

no hazard identified

Workers - Hazard for the eyes

Local effects

Hazard assessment conclusion:

low hazard (no threshold derived)

Additional information - workers

Relevant endpoint for DNEL derivation: toxicity to reproduction - NOAEL = 300 mg/kg bw/d

The study was performed according the requirements of OECD TG 422 guideline under GLP conditions. Following a previously conducted 14-day sighting study, the systemic toxic potential of the test item in rats, including a screen for reproductive/developmental effects and assessment of endocrine disruptor relevant endpoints was conducted by oral gavage administration for at least five weeks with additional subgroups used to assess reversibility, persistence or delayed effects for 14 days post treatment. Three toxicology treatment groups with a control was conducted, each comprising five or ten male and five female rats which received oral gavage test item at doses of 0 (Control), 150, 300 or 600 mg/kg bw/day test item formulated in corn oil vehicle. Ten males were treated in the 150 and 300 mg/kg bw/day doses for pairing purposes with the reproductive phase females. Recovery phase groups included five males and females treated at 0 (Control) and 600 mg/kg bw/day. Reproductive phase females, ten per group (10) were treated at doses of 0 (Control), 150, 300 or 600 mg/kg bw/day. Toxicity phase males were treated for two weeks before pairing up to necropsy after six weeks. Toxicity phase females were treated for six weeks. Recovery phase males were treated for two weeks before pairing up to necropsy after six weeks followed by a 2-week recovery period. Recovery phase females were treated for six weeks followed by a 2-week recovery period. Reproductive phase females were treated for two weeks before pairing, throughout pairing, gestation and until Day 12 of lactation. The offspring received no direct administration of the test item; any exposure was in utero or via the milk. Selected F1 offspring were sampled and killed on Day 4 or Day 13 of age for analysis of blood thyroid hormone levels. The remaining F1 offspring were killed on Day 13 of age. A similarly constituted Control group was assigned to each phase, and received, the vehicle: corn oil at the same dose volume as treated groups.

 

There were no treatment-related premature mortalities among adult animals during the course of the study. There was no adverse effect on clinical condition, sensory reactivity and grip strength, motor activity, ophthalmic changes and/or pre-coital interval, mating performance gestation length on the adult animals. Body weight gain was variable throughout the treatment period in males and non-mated females at 600 mg/kg bw/day. Overall gains were similar to control in both. Following cessation of treatment, females showed body weight loss, compared with body weight stasis in the Control. Body weight gain was variable during both gestation and lactation, but again overall gains in treated animals were similar to Control. Food consumption was similarly affected by treatment in all study phases, but was noticeably low during Days 4-10 of lactation. Water consumption was consistently high in males and non mated females at 300 or 600 mg/kg bw/day, during gestation at 600 mg/kg bw/day and lactation at 300 or 600 mg/kg bw/day. There were no toxicologically significant changes in haematology, blood chemistry and plasma parameters following five weeks of treatment in males and unmated females and on Day 13 of lactation except for minor elevated plasma cholesterol concentration in non-mated females and plasma urea concentration in males and minor change in creatinine concentrations in males/females receiving 600 mg/kg bw/day. These changes were attributed to changes in microscopic changes observed in male kidney and/or male/female liver. At the end of treatment liver weight and kidney weight were high in males/females at 300 and 600 mg/kg bw/day and 600 mg/kg bw/day, respectively. Urinalysis indicated a high urinary volume as a result of the higher water intake in males/females at 600 mg/kg bw/day. This may be attributed to higher plasma and urinary protein concentrations. Macroscopic examination performed after 5 weeks of treatment or 2 weeks recovery or reproductive females after day 13 lactation revealed no test item treatment related lesions. Microscopic examination revealed a dose-dependent hepatocellular periportal vacuolation in males and females treated at 150, 300 or 600 mg/kg/day that correlated at 600 mg/kg/day with increased storage of lipids within the cytoplasm of the hepatocytes and minor elevation of plasma cholesterol and elevated urea concentration in females or males in Week 5. This metabolic alteration may also be reflected by the presence of an increase in urinary ketones in all males at 600 mg/kg/day. The presence of periportal vacuolation correlated with the statistically significant increase in liver weights in males and females at 600 mg/kg/day. Examination of the liver in males and females at 600 mg/kg/day, following two weeks without treatment indicated partial recovery of the hepatocellular periportal vacuolation and normalisation of plasma cholesterol and urea concentrations. This treatment related change was not accompanied by inflammation or necrosis of the hepatocytes, therefore changes were considered unlikely to be adverse. A dose-dependent accumulation of hyaline droplets and basophilic tubules was evident in males receiving 300 or 600 mg/kg/day. Hyaline droplets are composed of poorly catabolized α2µ globulin probably binding to the test item and accumulating within the phagolysosomes of the renal tubular cells. The presence of increased α2µ globulin in males was confirmed immunohistochemically. Examination of the kidneys from Control and males previously treated at 600 mg/kg/day revealed resolution of the hyaline droplets however basophilic tubules persisted in the previously treated males. Basophilic tubules were only present focally and were consistent with background levels of this finding. Only partial recovery was evident following two weeks without treatment. Relevance for toxicity in humans through this α2µ globulin mechanism is considered unlikely.

 

At 600 mg/kg/day, two females were not pregnant. Three litters indicated mortality by Day 2, that was attributed to the reduced secretory activity of the mammary gland in the adults. Two females exhibited irregularities of the estrous cycle before mating. Of the three females with total litter loss, one showed extended estrus, and a second had an irregular cycle. The offspring from adults treated at 600 mg/kg/day were small on Day 1 of age and subsequent growth, especially during Days 4-7, to Day 13, correlated with clinical signs of poor maternal care. There was some evidence of poor maternal care at 300 mg/kg/day. Ano genital distance in the offspring was unaffected by maternal treatment. Male offspring did not develop nipples. T4 (thyroxine) levels were statistically low in males at 300 or 600 mg/kg bw/day but were comparable with control following two weeks without treatment. This change was reversible and with no pathological correlations was considered non-adverse.

 

Conclusion:

Under the conditions of this study, the no-observed-adverse-effect level (NOAEL) for systemic toxicity was considered to be 600 mg/kg bw/day. The α2µ globulin mediated kidney changes were deemed adverse, in the rat. However, toxicity to humans by this mechanism is considered unlikely. For reproductive / developmental toxicity for males and females a precautionary NOAEL is set at 300 mg/kg bw/day due to low pregnancy rate and high litter loss reported at 600 mg/kg bw/day. Applicant assessment indicates: that whilst the effects on lipid metabolism where not adverse in adults since they were deemed reversible, they may have led to adverse outcome in offspring via reduced secretory activity in female adults and consequential poor maternal care. On this basis a precautionary NOAEL has been adopted.

General Population - Hazard via inhalation route

Systemic effects

Long term exposure

Hazard assessment conclusion:

DNEL (Derived No Effect Level)

Value:

0.869 mg/m³

Most sensitive endpoint:

effect on fertility

Route of original study:

Oral

DNEL related information

DNEL derivation method:

ECHA REACH Guidance

Overall assessment factor (AF):

150

Dose descriptor starting point:

NOAEL

Value:

300 mg/kg bw/day

Modified dose descriptor starting point:

NOAEC

Value:

130.4 mg/m³

Explanation for the modification of the dose descriptor starting point:

Concerning absorption, in the absence of reliable data for both the starting route (oral) and the end route (inhalation), worst case assumptions were made. It was assumed that a limited absorption occurs by the oral route, leading to a low (conservative) internal NOAEL. To secure a conservative external NOAEL, a maximum absorption should be assumed for the inhalation route (i.e.; 100%) leading to a low external NOAEL. Thus, in the case of oral-to- inhalation extrapolation, it is proposed to include a default factor of 2, i.e. the absorption percentage by oral route is half that of the inhalation absorption as suggested in ECHA Guidance on Information Requirements and Chemical Safety Assessment, Chapter R.8 (2012).

To convert the oral NOAEL into inhalatory NOAEC, a rat default respiratory volume was used corresponding to the daily duration of human exposure (sRVrat: 1.15 m3/kg bw/24 h).

The corrected dose descriptor for inhalation is determined using the following equation:

Corrected Inhalator NOAEC = 1/sRVrat x ABSoral-rat/ABSinh-rat x ABSinh-rat/ABSinh-human

= [NOAEL] x [300 mg/kg bw/day] x [1/1.15 m3/kg bw/ day] x [1/2].

Thus, the corrected dose descriptor for inhalation is 130.4 mg/m3 for the general population.

AF for dose response relationship:

1

Justification:

The dose-descriptor is a NOAEL. Table R.8-6 ECHA REACH Guidance.

AF for differences in duration of exposure:

6

Justification:

Default factor for a sub-acute. Table R.8-5 ECHA REACH Guidance.

AF for interspecies differences (allometric scaling):

1

Justification:

Table R.8-4 ECHA REACH Guidance. Assessment factor not to be used for inhalation route since the differences in metabolic rate/bw has already been taken into account for the corrected dose descriptor.

AF for other interspecies differences:

2.5

Justification:

Default factor for other interspecies differences. Table R.8-6 ECHA REACH Guidance.

AF for intraspecies differences:

10

Justification:

Default factor for general population. Table R.8-6 ECHA REACH Guidance.

AF for the quality of the whole database:

1

Justification:

Default factor for good/standard quality of the database taken into account completeness of the standard information requirements for the tonnage band.

AF for remaining uncertainties:

1

Justification:

No remaining uncertainties.

Acute/short term exposure

Hazard assessment conclusion:

DNEL (Derived No Effect Level)

Value:

2.607 mg/m³

Most sensitive endpoint:

repeated dose toxicity

Route of original study:

Oral

DNEL related information

DNEL derivation method:

ECHA REACH Guidance

Overall assessment factor (AF):

150

DNEL extrapolated from long term DNEL

Dose descriptor starting point:

NOAEL

Value:

300 mg/kg bw/day

Modified dose descriptor starting point:

NOAEC

Value:

391.2 mg/m³

Explanation for the modification of the dose descriptor starting point:

Concerning absorption, in the absence of reliable data for both the starting route (oral) and the end route (inhalation), worst case assumptions were made. It was assumed that a limited absorption occurs by the oral route, leading to a low (conservative) internal NOAEL. To secure a conservative external NOAEL, a maximum absorption should be assumed for the inhalation route (i.e.; 100%) leading to a low external NOAEL. Thus, in the case of oral-to- inhalation extrapolation, it is proposed to include a default factor of 2, i.e. the absorption percentage by oral route is half that of the inhalation absorption as suggested in ECHA Guidance on Information Requirements and Chemical Safety Assessment, Chapter R.8 (2012).

To convert the oral NOAEL into inhalatory NOAEC, a rat default respiratory volume was used corresponding to the daily duration of human exposure (sRVrat: 1.15 m3/kg bw/24 h).

The corrected dose descriptor for inhalation is determined using the following equation:

Corrected Inhalator NOAEC = 1/sRVrat x ABSoral-rat/ABSinh-rat x ABSinh-rat/ABSinh-human

= [NOAEL] x [300 mg/kg bw/day] x [1/1.15 m3/kg bw/ day] x [1/2].

The extrapolation of long to short term risk was set based on a reference period of 15 minutes at 1-5 times the value (default 3) of the long-term DNEL i.e. Corrected Inhalatory NOAEC x 3.

Thus, the corrected dose descriptor for inhalation is 391.2 mg/m3 for workers.

AF for dose response relationship:

1

Justification:

The dose-descriptor is a NOAEL. Table R.8-6 ECHA REACH Guidance.

AF for interspecies differences (allometric scaling):

1

Justification:

Table R.8-4 ECHA REACH Guidance. Assessment factor not to be used for inhalation route since the differences in metabolic rate/bw has already been taken into account for the corrected dose descriptor.

AF for other interspecies differences:

2.5

Justification:

Default factor for other interspecies differences. Table R.8-6 ECHA REACH Guidance.

AF for intraspecies differences:

10

Justification:

Default factor for general population. Table R.8-6 ECHA REACH Guidance.

AF for the quality of the whole database:

1

Justification:

Default factor for good/standard quality of the database taken into account completeness of the standard information requirements for the tonnage band.

AF for remaining uncertainties:

6

Justification:

Default factor for a sub-acute. Table R.8-5 ECHA REACH Guidance.

Local effects

Long term exposure

Hazard assessment conclusion:

no hazard identified

Acute/short term exposure

Hazard assessment conclusion:

no hazard identified

DNEL related information

General Population - Hazard via dermal route

Systemic effects

Long term exposure

Hazard assessment conclusion:

DNEL (Derived No Effect Level)

Value:

0.5 mg/kg bw/day

Most sensitive endpoint:

effect on fertility

Route of original study:

Oral

DNEL related information

DNEL derivation method:

ECHA REACH Guidance

Overall assessment factor (AF):

600

Dose descriptor starting point:

NOAEL

Value:

300 mg/kg bw/day

Value:

300 mg/kg bw/day

Explanation for the modification of the dose descriptor starting point:

For systemic hazard assesment via the dermal route of exposure, route-to-route extrapolation from the oral NOAEL value was considered appropriate. As no data on dermal penetration are available, dermal absorption is considered to be the same as oral absorption (i.e. 100 %). Therefore the oral NOAEL is considered the same as the dermal NOAEL (ECHA Guidance on Information Requirements and Chemical Safety Assessment, Chapter R.8 (2012)).

AF for dose response relationship:

1

Justification:

The dose-descriptor is a NOAEL. Table R.8-6 ECHA REACH Guidance.

AF for differences in duration of exposure:

6

Justification:

Default factor for a sub-acute study. Table R.8-5 ECHA REACH Guidance.

AF for interspecies differences (allometric scaling):

4

Justification:

Default allometric scaling factor for rats. Table R.8-4 ECHA REACH Guidance.

AF for other interspecies differences:

2.5

Justification:

Default factor for other interspecies differences. Table R.8-6 ECHA REACH Guidance.

AF for intraspecies differences:

10

Justification:

Default factor for general population. Table R.8-6 ECHA REACH Guidance.

AF for the quality of the whole database:

1

Justification:

Default factor for good/standard quality of the database taken into account completeness of the standard information requirements for the tonnage band.

AF for remaining uncertainties:

1

Justification:

No remaining uncertainties.

Acute/short term exposure

Hazard assessment conclusion:

no hazard identified

DNEL related information

Local effects

Long term exposure

Hazard assessment conclusion:

no hazard identified

Acute/short term exposure

Hazard assessment conclusion:

no hazard identified

General Population - Hazard via oral route

Systemic effects

Long term exposure

Hazard assessment conclusion:

DNEL (Derived No Effect Level)

Value:

0.5 mg/kg bw/day

Most sensitive endpoint:

effect on fertility

Route of original study:

Oral

DNEL related information

DNEL derivation method:

ECHA REACH Guidance

Overall assessment factor (AF):

600

Dose descriptor starting point:

NOAEL

Value:

300 mg/kg bw/day

Value:

300 mg/kg bw/day

Explanation for the modification of the dose descriptor starting point:

No modification of the dose descriptor starting point is required. The endpoint used to derive the DNEL uses the oral route for exposure.

AF for dose response relationship:

1

Justification:

The dose-descriptor is a NOAEL. Table R.8-6 ECHA REACH Guidance.

AF for differences in duration of exposure:

6

Justification:

Default factor for a sub-acute study. Table R.8-5 ECHA REACH Guidance.

AF for interspecies differences (allometric scaling):

4

Justification:

Default allometric scaling factor for rats. Table R.8-4 ECHA REACH Guidance.

AF for other interspecies differences:

2.5

Justification:

Default factor for other interspecies differences. Table R.8-6 ECHA REACH Guidance.

AF for intraspecies differences:

10

Justification:

Default factor for general population. Table R.8-6 ECHA REACH Guidance.

AF for the quality of the whole database:

1

Justification:

Default factor for good/standard quality of the database taken into account completeness of the standard information requirements for the tonnage band.

AF for remaining uncertainties:

1

Justification:

No remaining uncertainties.

Acute/short term exposure

Hazard assessment conclusion:

no hazard identified

DNEL related information

General Population - Hazard for the eyes

Local effects

Hazard assessment conclusion:

low hazard (no threshold derived)

Additional information - General Population

Relevant endpoint for DNEL derivation: toxicity to reproduction - NOAEL = 300 mg/kg bw/d

The study was performed according the requirements of OECD TG 422 guideline under GLP conditions. Following a previously conducted 14-day sighting study, the systemic toxic potential of the test item in rats, including a screen for reproductive/developmental effects and assessment of endocrine disruptor relevant endpoints was conducted by oral gavage administration for at least five weeks with additional subgroups used to assess reversibility, persistence or delayed effects for 14 days post treatment. Three toxicology treatment groups with a control was conducted, each comprising five or ten male and five female rats which received oral gavage test item at doses of 0 (Control), 150, 300 or 600 mg/kg bw/day test item formulated in corn oil vehicle. Ten males were treated in the 150 and 300 mg/kg bw/day doses for pairing purposes with the reproductive phase females. Recovery phase groups included five males and females treated at 0 (Control) and 600 mg/kg bw/day. Reproductive phase females, ten per group (10) were treated at doses of 0 (Control), 150, 300 or 600 mg/kg bw/day. Toxicity phase males were treated for two weeks before pairing up to necropsy after six weeks. Toxicity phase females were treated for six weeks. Recovery phase males were treated for two weeks before pairing up to necropsy after six weeks followed by a 2-week recovery period. Recovery phase females were treated for six weeks followed by a 2-week recovery period. Reproductive phase females were treated for two weeks before pairing, throughout pairing, gestation and until Day 12 of lactation. The offspring received no direct administration of the test item; any exposure was in utero or via the milk. Selected F1 offspring were sampled and killed on Day 4 or Day 13 of age for analysis of blood thyroid hormone levels. The remaining F1 offspring were killed on Day 13 of age. A similarly constituted Control group was assigned to each phase, and received, the vehicle: corn oil at the same dose volume as treated groups.

 

There were no treatment-related premature mortalities among adult animals during the course of the study. There was no adverse effect on clinical condition, sensory reactivity and grip strength, motor activity, ophthalmic changes and/or pre-coital interval, mating performance gestation length on the adult animals. Body weight gain was variable throughout the treatment period in males and non-mated females at 600 mg/kg bw/day. Overall gains were similar to control in both. Following cessation of treatment, females showed body weight loss, compared with body weight stasis in the Control. Body weight gain was variable during both gestation and lactation, but again overall gains in treated animals were similar to Control. Food consumption was similarly affected by treatment in all study phases, but was noticeably low during Days 4-10 of lactation. Water consumption was consistently high in males and non mated females at 300 or 600 mg/kg bw/day, during gestation at 600 mg/kg bw/day and lactation at 300 or 600 mg/kg bw/day. There were no toxicologically significant changes in haematology, blood chemistry and plasma parameters following five weeks of treatment in males and unmated females and on Day 13 of lactation except for minor elevated plasma cholesterol concentration in non-mated females and plasma urea concentration in males and minor change in creatinine concentrations in males/females receiving 600 mg/kg bw/day. These changes were attributed to changes in microscopic changes observed in male kidney and/or male/female liver. At the end of treatment liver weight and kidney weight were high in males/females at 300 and 600 mg/kg bw/day and 600 mg/kg bw/day, respectively. Urinalysis indicated a high urinary volume as a result of the higher water intake in males/females at 600 mg/kg bw/day. This may be attributed to higher plasma and urinary protein concentrations. Macroscopic examination performed after 5 weeks of treatment or 2 weeks recovery or reproductive females after day 13 lactation revealed no test item treatment related lesions. Microscopic examination revealed a dose-dependent hepatocellular periportal vacuolation in males and females treated at 150, 300 or 600 mg/kg/day that correlated at 600 mg/kg/day with increased storage of lipids within the cytoplasm of the hepatocytes and minor elevation of plasma cholesterol and elevated urea concentration in females or males in Week 5. This metabolic alteration may also be reflected by the presence of an increase in urinary ketones in all males at 600 mg/kg/day. The presence of periportal vacuolation correlated with the statistically significant increase in liver weights in males and females at 600 mg/kg/day. Examination of the liver in males and females at 600 mg/kg/day, following two weeks without treatment indicated partial recovery of the hepatocellular periportal vacuolation and normalisation of plasma cholesterol and urea concentrations. This treatment related change was not accompanied by inflammation or necrosis of the hepatocytes, therefore changes were considered unlikely to be adverse. A dose-dependent accumulation of hyaline droplets and basophilic tubules was evident in males receiving 300 or 600 mg/kg/day. Hyaline droplets are composed of poorly catabolized α2µ globulin probably binding to the test item and accumulating within the phagolysosomes of the renal tubular cells. The presence of increased α2µ globulin in males was confirmed immunohistochemically. Examination of the kidneys from Control and males previously treated at 600 mg/kg/day revealed resolution of the hyaline droplets however basophilic tubules persisted in the previously treated males. Basophilic tubules were only present focally and were consistent with background levels of this finding. Only partial recovery was evident following two weeks without treatment. Relevance for toxicity in humans through this α2µ globulin mechanism is considered unlikely.

 

At 600 mg/kg/day, two females were not pregnant. Three litters indicated mortality by Day 2, that was attributed to the reduced secretory activity of the mammary gland in the adults. Two females exhibited irregularities of the estrous cycle before mating. Of the three females with total litter loss, one showed extended estrus, and a second had an irregular cycle. The offspring from adults treated at 600 mg/kg/day were small on Day 1 of age and subsequent growth, especially during Days 4-7, to Day 13, correlated with clinical signs of poor maternal care. There was some evidence of poor maternal care at 300 mg/kg/day. Ano genital distance in the offspring was unaffected by maternal treatment. Male offspring did not develop nipples. T4 (thyroxine) levels were statistically low in males at 300 or 600 mg/kg bw/day but were comparable with control following two weeks without treatment. This change was reversible and with no pathological correlations was considered non-adverse.

 

Conclusion:

Under the conditions of this study, the no-observed-adverse-effect level (NOAEL) for systemic toxicity was considered to be 600 mg/kg bw/day. The α2µ globulin mediated kidney changes were deemed adverse, in the rat. However, toxicity to humans by this mechanism is considered unlikely. For reproductive / developmental toxicity for males and females a precautionary NOAEL is set at 300 mg/kg bw/day due to low pregnancy rate and high litter loss reported at 600 mg/kg bw/day. Applicant assessment indicates: that whilst the effects on lipid metabolism where not adverse in adults since they were deemed reversible, they may have led to adverse outcome in offspring via reduced secretory activity in female adults and consequential poor maternal care. On this basis a precautionary NOAEL has been adopted.