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Description of key information

Toxicokinetic data demonstrate very low or absent systemic bioavailability. This is consistent with results from a key, GLP compliant guideline screening study which found no systemic effects following repeated oral exposure. Results from a key GLP compliant guideline sub-chronic (90-d) inhalation study identify a NOAEC (threshold) for hyperplasia in nasal tissue.

Key value for chemical safety assessment

Repeated dose toxicity: inhalation - local effects

Endpoint conclusion
Endpoint conclusion:
adverse effect observed
Dose descriptor:
NOAEC
22.2 mg/m³
Study duration:
subchronic
Species:
rat

Additional information

All studies reported in this section were conducted using TBHP (aqueous 70% solution, as manufactured and supplied): results have been expressed in terms of dry TBHP.

ORAL

Results are available from a combined repeated dose and reproductive/developmental toxicity screening test (OECD 422) where TBHP was administered by oral gavage to male and female Wistar rats at received doses of up to 21 mg/kg bwt/d for up to 45 days (TNO Study Director, 1993). The study was GLP-compliant, with dose level selection based on the results of two preliminary range-finding investigations which showed moderate to severe damage to the stomach mucosa following administration of doses of 35 mg TBHP/kg bwt/day and above for 5 days. A dose-related increase in the incidence of renal lesions (described by the study pathologist as multifocal increased accumulation of tubular proteinaceous material and multifocal tubular nephrosis) was observed in males at all doses (females unaffected). There were no other findings of note. The nominal systemic NOEL for this finding in male rats was 2.1 mg TBHP/kg bwt/day; the nominal NOAEL for systemic toxicity (both sexes) was 21 mg/kg bwt/d (the highest dose tested).

 

A subsequent mechanistic investigation (TNO Study Director, 2002) also demonstrated the occurence of mild kidney changes (described by the study pathologist as very slight to slight hyaline droplet nephropathy) in a majority of male rats given 21 mg TBHP/kg bwt for 42 days but not in controls. A positive immunohistochemical response for alpha-2u-globulin was found for all control and all test kidneys although the response in a majority of the TBHP-treated animals tended to be more intense than that seen in the controls. These results are therefore inconclusive with regard to the possible involvement of alpha-2u-globulin in the kidney changes observed in this study and also in the OECD 422 investigation.

 

A retrospective expert histological evaluation of kidney tissue from the two preceding studies (Hard, 2007) did not confirm the induction of hyaline droplets or any renal changes associated with alpha-2u-globulin nephropathy; nor did TBHP exacerbate the occurrence of chronic progressive nephropathy. The report concluded that TBHP caused no nephropathic alteration in either study. This conclusion was endorsed by the EU Risk assessment Report for TBHP (document reference R319_0710_HH, RIVM Chemical Substances Bureau, version dated November 2007). The overall systemic NOAEL for TBHP after repeated oral exposure was therefore 21 mg/kg bwt/d.

 

Mechanistic metabolic data demonstrate that the systemic bioavailability of TBHP is very low or absent due to its rapid conversion to 2-methylpropan-2-ol. Repeated dose oral toxicity studies on 2-methylpropan-2-ol show an interaction with alpha-2u-globulin in male rat kidney whereas no such interaction was found in the available studies on TBHP. This may reflect differences in treatment levels between the studies, with doses tested for TBHP (21 mg/kg bwt/day) being lower than those used with 2-methylpropan-2-ol (up to 200 mg/kg bwt/day). It is noted that exposure to higher doses of TBHP is not practical or reasonable given the corrosive nature of the substance. This is confirmed by the occurrence of moderate to severe stomach damage reported in rats following sub-acute gavage administration at 35 mg/kg bwt/day (TNO Study Director, 1993).

 

INHALATION

Sub-Acute Toxicity

The sub-acute inhalation toxicity of TBHP was investigated in groups of male and female Wistar rats exposed head-nose only to 0, 2, 6 or 18 ppm vapor (0, 7.4, 22.2 or 66.7 mg/m3) 6 hr/d, 5 d/wk for 4 wk (BASF, 2009). Following the last exposure, blood was sampled and clinical chemistry and haematology parameters determined using standard methods. The animals were then subject to gross necropsy, including macroscopic examination of the major internal organs and collection of organ weight data. Selected tissues were processed and examined microscopically with particular emphasis on the nasal cavity (4 levels) and larynx (3 levels). Determinations of cell proliferation (BrdU incorporation) were also performed on nasal tissue. The investigation was GLP compliant and followed OECD guideline 412.

All animals survived to scheduled necropsy with no treatment-related clinical observations. Body weight and body weight gain were decreased in high dose males on exposure days 7 and 14 (possibly reflecting a transitory response to the irritative properties of TBHP vapor) but were similar to control values thereafter. Findings at gross necropsy were unremarkable, however histopathological assessment treatment-related alterations in the nasal cavity from high dose males and females comprising (i) minimal to mild increased thickness of the transitional epithelium and (ii) slight hyperplasia / metaplasia (e.g. reflecting a change towards squamous epithelium). The tip of the maxillary turbinate and the lateral wall of the nasal cavity were mainly affected while other levels of the nasal cavity were free of lesions. Microscopic findings in other tissues examined (e.g. larynx, mediastinal lymph nodes, traecheobronchial lymph nodes, trachea, lung, liver, kidneys) were spontaneous in origin. Cell proliferation determinations revealed significantly increased unit length labeling indices in transitional epithelium lining Area 1 (maxillary turbinate) and Area 2 (lateral wall) from level I of the nasal cavity in high dose males and females i.e. responses present to those sites exhibiting histopathological lesions. No increased cell proliferation was detected in levels II-IV of the nasal cavity.

The results demonstrate that Inhalation exposure to TBHP vapour for 28 days resulted in hyperplasia / metaplasia of the transitional epithelia of the nasal cavity with an associated increased S-phase response (the latter present only in tissue exhibiting microscopic changes). Based on these findings, the NOAEC for local toxicity of TBHP following inhalation exposure was 6 ppm (22.2 mg/m3).

Sub-Chronic Toxicity

The sub-chronic inhalation toxicity of TBHP was investigated in groups of male and female Wistar rats exposed head-nose only to 0, 2, 6 or 18 ppm vapor (0, 7.4, 22.2 or 66.7 mg/m3) 6 hr/d, 5 d/wk for 13 wk (WIL, 2013). Following the last exposure, blood was sampled and clinical chemistry and haematology parameters determined using standard methods. The animals were then subject to gross necropsy, including macroscopic examination of the major internal organs and collection of organ weight data. Selected tissues were processed and examined microscopically with particular emphasis on the nasal cavity (6 levels). The investigation was GLP compliant and followed OECD guideline 413.

All animals survived to scheduled necropsy with no treatment-related clinical observations. Findings at gross necropsy were unremarkable, however histopathological assessment treatment-related alterations in the nasal cavity from high dose males and females comprising transitional epithelium hyperplasia characterized by a minimal to mild increase in the thickness of the transitional epithelium (e.g. reflecting a change towards squamous epithelium). The hyperplastic regions were predominantly located at the tips of the maxilloturbinates and nasoturbinates and along the lateral wall of the nasal cavity. Acute inflammation in the nasal cavity was also observed, although it was not considered to be test substance related. There were no other test substance related histologic changes. Hyperplasia was only noted in one post-exposure recovery female and that was considered secondary to the inflammation in the animal rather than related to administration of the test substance.

The results demonstrate that Inhalation exposure to TBHP vapour for 90 days resulted in a temporal hyperplasia / metaplasia of the transitional epithelia of the nasal cavity. Based on these findings, the NOAEC for local toxicity of TBHP following inhalation exposure was 6 ppm (22.2 mg/m3).

Justification for classification or non-classification

Toxicokinetic data demonstrate rapid conversion of TBHP to TBA in vivo limiting the potential for systemic exposure. Morbidity seen in repeated dose oral studies is presumed to occur secondary to site of contact corrosion of the gastric mucosae. TBHP is corrosive to biological tissue, indicating that local (site of contact) effects are likely, as confirmed by results from a sub-chronic (90 day) inhalation toxicity study.

The 90 -day study also provides a NOAEC for subtle cell proliferative and histopathological changes that were temporal in nature.

No classification required under 67/548 EEC, the CLP regulation or UN GHS criteria.