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Description of key information

In repeated dose studies, the principle effects of xylenes were adaptive changes in the liver, changes in kidney and liver weights,  body weight changes and minimal nephropathy in females.  

Key value for chemical safety assessment

Repeated dose toxicity: via oral route - systemic effects

Endpoint conclusion
Dose descriptor:
250 mg/kg bw/day

Repeated dose toxicity: inhalation - systemic effects

Endpoint conclusion
Dose descriptor:
3 515 mg/m³

Additional information

Mixed xylene (CAS 1330-20-7) comprises individual xylene isomers (m-xylene, o-xylene, p-xylene) and ethylbenzene. Data for these substances and the specific component substances benzene, toluene and styrene have been included as supporting information.

Repeated dose toxicity: oral

No studies are available for the individual xylene isomers.

Data are available for laboratory animals exposed to high doses of mixed xylenes, adverse effects have been observed in the kidney and liver (IARC, 1989).

A near-guideline (equivalent or similar to OECD 408) subchronic oral gavage study with mixed xylenes (20% ethylbenzene) was conducted by Condie et al. (1988). It includes the range of toxicological endpoints routinely investigated in regulatory subchronic studies and is a key study for identifying the effects of mixed xylenes. In the study, groups of 10 male and 10 female rats were given 0, 150, 750, or 1500 mg/kg bw/day of mixed xylenes in corn oil for 90 consecutive days. A decrease in body weight was observed in males only at 1500 mg/kg bw/day. Although increased relative liver weights were seen at all dose levels in males and in females at 750 and 1500 mg/kg bw/day there were no adverse histopathology findings in the liver. An increased relative kidney weight was observed in high dose males and females and in intermediate dose males. In males there was a dose-related increase in the incidence of slight to mild hyaline droplet formation in tubules at all dose levels. This finding is indicative of alpha-2u-globulin which is considered to be male rat-specific and is not relevant for humans. In females the incidence of minimal nephropathy in females was statistically significantly increased in the 750 and 1500 mg/kg bw/day groups. This finding described as scattered tubular dilation and atrophy, with occasional regeneration and is the chronic progressive nephropathy typically seen in ageing rats.

No NOAEL was established in this study for males based on liver weight increases. However increases in liver weight with no adverse histopathological findings are considered to be an adaptive response to administration of mixed xylenes rather than an adverse toxicological effect. A dose of 750 mg/kg bw/day is the NOAEL based on effects on male body weight. In females a NOAEL of 150 mg/kg bw/day is based on liver weight increases and the kidney effects observed at dose levels of 750 mg/kg bw/day and higher.

Treatment levels between 150 and 750 mg/kg bw/day are covered in a carcinogenicity study in rats (NTP, 1986). Although this study did not include all of the end points included in chronic studies to current guidelines, the key parameters affected in the sub chronic study, i.e. body weights and detailed pathology and histopathology are included. Rats were dosed with mixed xylenes at concentrations of 0, 250 or 500 mg/kg bw/day 5 days per week for 103 weeks. The main finding was a decrease in body weights in males receiving 500 mg/kg bw/day in the second year of the study.There was no other evidence of systemic toxicity including no treatment-related pathology findings. A dose of 250 mg/kg/day was a NOAEL for both sexes and this is considered to be the key study for determining the NOAEL for repeated dose exposure to mixed xylenes via the oral route.

A supporting subchronic oral gavage study was conducted by NTP (1986) and included both rats and mice. The toxicity of mixed xylenes in corn oil was investigated in groups of 10 male and 10 female rats following gavage dosing on 5days/week for 13 weeks with 0, 62.5, 125, 250, 500 or 1000 mg/kg/day. In the same study, groups of 10 male and 10 female mice were similarly dosed with 0, 125, 250, 500, 1000 or 2000 mg/kg/day of mixed xylenes in corn oil. Limited toxicological endpoints were evaluated.

Treatment-related findings in rats were limited to a reduction in overall body weight gain (15% for males and 8% for females) with a NOAEL of 500 mg/kg/day. High dose mice exhibited transient CNS effects 5-10 minutes after dosing that lasted 15-60 minutes. Other treatment-related findings were limited to a reduction in overall body weight gain (7% for males and 17% for females) with a NOAEL of 1000 mg/kg/day. Neither blood clinical chemistry nor organ weight data were collected for either species. No treatment-related macroscopic or microscopic lesions were observed in the tissues examined including the liver and kidney.

In an OECD Guideline 90-day oral study ethylbenzene was gavage dosed at 0, 75, 250 and 750 mg/kg in corn oil (Mellert et al. 2007). The NOAEL for this study was 75 mg/kg/day based on changes in haematology indicative of a mild regenerative anaemia and changes in clinical chemistry parameters. There was also an increase in liver weights with centrilobular hepatocellular hypertrophy indicative of hepatic microsomal enzyme induction.

Repeated dose toxicity: dermal

No studies are available for mixed xylene, the individual xylene isomers or ethylbenzene.

Repeated dose toxicity: inhalation

The subchronic inhalation studies are designed primarily to address neurological endpoints in male rats and dogs. These endpoints are discussed in section

In a recent study by Gagnaire et al (2001) the potential ototoxicity of individual xylene isomers was evaluated using electrophysiological methods in male rats exposed by inhalation to three different concentrations 6 hours/day, 5 days/week for 13 weeks was evaluated. The highest exposure concentration of 1800 ppm had no significant effect on body weight or body weight gain.

Mixed xylenes were assessed following a comparable protocol (Gagnaire et al, 2007a). Groups of male rats were exposed to 250, 500, 1000 and 2000 ppm mixed xylenes, one mixture with 10% and one with 20% ethylbenzene for 6 h/day, 6 d/wk over 13 weeks with a recovery period of 8 weeks. There was no adverse effect on body weight at any of the dose levels.

In an older study by Carpenter (1975), male rats and male dogs were exposed 6h/day for 5 days in each of 13 weeks to 0, 180, 460 or 810 ppm mixed xylenes. The highest exposure level was a NOAEC for both species.

Ethylbenzene (Not currently classified): No repeated dose toxicity studies in humans have been identified. The EU transitional RAR (EU, 2008b) concluded "Repeat-dose or prolonged exposure to ethylbenzene specifically affected the nervous system, but did not induce overt toxicity of any other organ system. " The auditory system is the most sensitive to the toxic effects of ethylbenzene after inhalation exposure (Gagnaire et al, 2007) while the liver is the most sensitive following oral exposure (Mellert et al, 2006). The LOAEL for ototoxicity was 200 ppm (868 mg/m3) and the NOAEL for hepatotoxicity was 75 mg/kg/day in a 13 week oral gavage study in rats.

Other specific components which have been identified as potentially present in some streams are styrene, benzene and toluene. These are all identified as producing serious target organ toxicity following repeated oral, dermal or inhalation exposures in animals and man.

Styrene (Not currently classified): Inhalation studies in animals have reported damage to the nasal olfactory epithelium in rats and mice, liver damage in mice, eye irritation in rats and guinea pigs, ototoxicity in rats and impaired nerve conduction velocity. The EU transitional RAR (EU, 2008b) has identified a NOAEC of 50 ppm in humans based on colour vision discrimination effects in occupationally exposed workers.

Benzene (Classification: EU -Toxic T, R48/23/24/25; GHS/CLP - STOT-RE Category 1, H372): After repeated dose exposure via oral or inhalation routes, benzene causes adverse effects on the haematopoietic system of animals and man. The oral LOAEL was 25 mg/kg bw/day for male and female mice (NTP, 1986) and the inhalation LOAEC for haematotoxicity in mice is 10 ppm (32 mg/m3) (Ward et al, 1985). For human a NOAEC of 3.5 ppm (11.2 mg/m3) is obtained based on the 95% LCL for the threshold level of neutrophils, the most sensitive endpoint reported by Schnatter et al. (2010).

Toluene (Classification: EU - Harmful Xn, R48/20; GHS/CLP - STOT-RE Category 2, H373): Toluene exposure can produce central nervous system pathology in animals after high oral doses. Repeated inhalation exposure can produce ototoxicity in the rat and high concentrations are associated with local toxicity (nasal erosion). In humans neuropsychological effects and disturbances of auditory function and colour vision have been reported, particularly when exposures are not well controlled and/or associated with noisy environments. The NOAEC for subchronic oral toxicity in rats is 625 mg/kg/day based on neuropathology (Huff, 1990). The NOAEC for inhalation toxicity in the rat is 300 ppm (1131 mg/m3) based on effects on body weight, mortality and adverse local effects (nasal erosion) (Gibson and Hardisty, 1983). The NOAEC for neuropsychological effects, auditory dysfunction and disturbances of colour vision in humans is 26 ppm (98 mg/m3) (Seeber et al, 2004); Schaper et al, 2003, 2004).


ACC, (2005). US High Production Chemical Program. Category summary for Fuel Oils Category. American Chemical Council. http: //www. epa. gov/chemrtk/pubs/summaries/fueloils/c13435tc. htm

Cragg ST, Clarke EA, Daly IW, Miller RR, Terrill JB and Quellette RE(1989). Subchronic inhalation toxicity of ethylbenzene in mice, rats, and rabbits. Fundam Appl Toxicol 13, 399-408.

NTP (1992). Toxicity studies of ethylbenzene in F344/N rats and B6C3F1 mice (inhalation studies). Research Triangle Park, NC: National Toxicology Program. NTP Tox 10. http: //inmagic. syrres. com/esc-images/NTIS/NTP/TOX010. pdf. August 02, 2007.


Justification for classification or non-classification

Mixed xylenes are not classified for repeat dose toxicity. However specific components may trigger classification. The maximum percentage of benzene is 1% and streams which contain 1% benzene are expected to cause adverse effects and should be classified as Harmful Xn R48/20/21/22 according to Dir 67/548/EEC and Cat 2, H373 according to GHS/CLP. Although ethylbenzene and styrene are not classified, they are expected to cause adverse effects. Mixed xylenes where the concentration of ethylbenzene or styrene is ≥10% should be self classified under DSD or CLP as Xn, R48/20 with the equivalent classification as STOT-RE Cat 2 H373 under CLP (see Specific Investigations: other studies (ototoxicity). The maximum concentration of toluene present in mixed xylenes is below the level which would trigger classification.