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Carcinogenicity

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Description of key information

Carcinogenicity tests have not been conducted on 1,2,4-trimethylbenzene or other trimethylbenzene isomers but there are sufficient data to indicate that they do not pose a significant carcinogenic hazard to humans. Trimethylbenzenes have been shown to have no significant genotoxic activity in vitro and in vivo. There are no data from toxicokinetics, metabolism and distribution findings to indicate any target for possible genotoxic carcinogenic activity. The cellular changes arising from repeated administration of trimethylbenzenes to animals are also minimal.

Key value for chemical safety assessment

Additional information

This endpoint summary provides an evaluation covering the range of trimethylbenzene materials including 1,2,3-trimethylbenzene, 1,2,4-trimethylbenzene and 1,3,5-trimethylbenzene.

There are no lifetime animal studies available on trimethylbenzene in order to directly investigate for possible carcinogenic activity. However, there is sufficient information available to make a satisfactory assessment of the likely carcinogenic potential of the material.

a) Non-testing information relevant for carcinogenicity

Trimethylbenzene is a simple aliphatic structure. It contains no centres of chemical functional activity and contains no alerts for possible genotoxic activity when considered against the criteria established by Ashby and Tennant (1988). The chemical would therefore not be expected to show any significant genotoxic activity in vitro or in vivo, and therefore would also not be expected to show carcinogenic activity due to a genotoxic mechanism. 

The metabolism and disposition of 1,2,4-trimethylbenzene and 1,3,5-trimethylbenzene have been examined in animals and are quite similar. Both materials form dimethylhippuric acid in rats administered a single oral dose. Inhalation toxicokinetic studies in humans show similar elimination kinetics and metabolism. The major route of metabolism is therefore through oxidation and subsequent conjugation of the methyl groups. None of these chemicals carry any alerts for possible genotoxic activity when considered as above.

Trimethylbenzenes are therefore relatively simple aromatic chemicals, which, when evaluated using accepted structure activity relationship considerations, have no alerts for significant genotoxicity or subsequent carcinogenic activity.

b) Testing data relevant for carcinogenicity

In vitro data

Trimethylbenzene isomers have been examined for mutagenicity in vitro in a range of recognised core assay types, including in bacteria (Ames test for gene mutation), and in mammalian cells (gene mutation, chromosomal damage and sister chromatid exchange endpoints).  Negative results were obtained in all assays, except for 1,2,3-trimethylbenzene, which gave a positive response in bacteria only, and then only in the absence of a mammalian metabolising system. In the presence of a mammalian metabolising system it gave a negative response.  Overall, it is concluded that the data do not indicate any significant genotoxic activity for 1,2,4-trimethylbenzene. 

Animal data

a) Genotoxicity studies

Trimethylbenzene isomers have been examined in the mouse bone marrow micronucleus assay, and were found not to be mutagenic. This is an established core assay for the evaluation of mutagenicity in vivo and indicative of no mutagenic activity in animals. In contrast, the reporting of data in the same study for the endpoint of sister chromatid exchange is not considered to be an observation of such weight in an evaluation of mutagenicity. The observation of small increases in sister chromatid exchange in the mouse for the high flash naptha (containing 55% trimethylbenzene isomers) is therefore not considered to indicate a significant genotoxic effect, since the increases were small, and the endpoint is one of uncertain relevance.

It is concluded, based on a range of in vitro and in vivo studies that the available data indicates that 1,2,4-trimethylbenzene does not have no significant genotoxicity. There is therefore good evidence for 1,2,4-trimethylbenzene being non-carcinogenic in animals with respect to a genotoxic mechanism.

b) Toxicity studies

A number of repeat dose toxicity studies in animals of up to 90 days duration are available using TMB isomers or mixtures containing TMB isomers. Studies have been undertaken using both the oral (gavage) and inhalation routes.

 

Gavage

 

1,2,3 -Trimethylbenzene and 1,2,4-trimethylbenzene were examined in 28 day oral gavage studies in rats at doses up to 1000 mg/kg/day.  For 1,2,3-trimethylbenzene, increases in liver and kidney weights were observed together with decreased thymus weights, clinical chemistry changes and at the top dose only, hyaline droplets and eosinophilic changes in the kidneys of male animals. For 1,2,4-trimethylbenzene, increases in liver and kidney weights were again observed, but no haematological changes or abnormalities at necropsy. A 90 day oral gavage study in rats was conducted with 1,3,5-trimethylbenzene, at doses up to 600 mg/kg/day, including a recovery group terminated after a further 28 day period. The changes reported were increased liver weights, increased kidney weights (males only), and increases serum phosphorus levels. No microscopic changes related to treatment were observed in the liver, kidney or other organs. No effects were observed in animals at the end of the 28 day recovery period, and all of the effects seen after 90 days were considered adaptive changes.

Inhalation

1,2,3 -Trimethylbenzene administered to rats by inhalation at dose levels up to 1230 mg/m3 for 6h/day, 5 days/week for 90 days resulted in increased liver weights in males and some limited clinical chemistry changes in both sexes. Rats were exposed to 1,2,4-trimethylbenzene by inhalation at dose levels up to 1250 mg/m3 for 6h/day, 5 days/week for 90 days, and no toxicologically significant changes were reported.  A second 90 day inhalation study in the rat reported only minimal clinical chemistry changes that were considered to be adaptive in nature. 1,3,5-trimethylbenzene was administered to rats by inhalation for 6h/day, 5 days/week for 5 weeks at 3050 mg/m3. Limited changes in white blood cells were the only effects reported. 

Exposure of rats by inhalation to a C9-C10 fraction from petroleum distillates, containing 74% alkylbenzenes (Firth, 2008), at a level of 616ppm for 135 days gave rise to congestive and hemorrhagic changes in lungs, kidneys, spleen and omental tissue. Inflammatory changes in the lung, and haemorrhages from the mouth and nose and transitory blood changes were also reported. 

Exposure of rats by inhalation for 13 weeks to a 50:50 blend of Shellsol A and Solvesso 100 (a C9-C10 dialkyl and trialkylbenzene mixture) up to 1800 mg/m3, resulted in increased liver and kidney weights in males, but no histopathological changes. The changes were considered adaptive in nature and a NOAEC of 1800 mg/m3 was reported.

The liver and kidney are the two main organs that showed some changes following exposure to trimethylbenzenes.  In both cases increased liver weights were seen. No histopathological changes were observed in the liver, and the minimal changes seen at 90 days were reversible after a recovery period of a further 28 days. This is consistent with a common reaction of the liver to continued exposure to a chemical that undergoes hepatic metabolism, and is considered an adaptive change. The only histopathological changes reported in the kidney on repeated exposure were hyaline droplets and eosinophilic bodies observed only after 1,2,3-TMB administration. These changes were only reported in male animals, and are consistent with the formation of the alpha-2u globulin protein. This is a relatively common response following exposure of male rats to certain hydrocarbons. The phenomenon has been well studied and it has been shown that it is a response specific to male rats, and it is generally agreed that it is an observation of no relevance to humans (USEPA 1991). 

It is possible that the underlying modes of action causing these changes in the liver and kidney may, on exposure of appropriate rodents to high levels of TMBs for the duration of a lifetime, result in an increase in tumours in the liver and kidney. Any tumours in the kidney would be expected in male rats only. If such tumours were observed, they would be considered to have arisen by established and accepted non-genotoxic modes of action, and would be appropriate for assessment by using standard toxicological assessment, and not by any assessment for genotoxic carcinogenicity. Any kidney tumours that may be induced would be expected to be in male rats and due to a mechanism involving alpha-2u globulin protein and would be considered to have no relevance for humans.

The other various changes in clinical chemistry or blood parameters are not considered to indicate changes or mechanisms likely to induce tumours on chronic exposure. 

Various trimethylbenzene isomers, either as pure chemicals or as components of mixtures have been examined in a range of studies of approximately 90 days including both oral and inhalation routes of exposure. The effects produced have been described including histopathological evaluation, and including an evaluation after a recovery period for the 1,3,5-TMB isomer. The effects reported have been relatively limited and are sufficient to allow an assessment of the likely effects of chronic exposure to TMBs.  It is considered unlikely that significant new data would be gained by undertaking specific chronic exposure studies.

The toxicokinetics, metabolism and distribution findings for 1,2,4-TMB and other isomers in animals indicate a chemical that is cleared primarily and rapidly through the urine, and with no accumulation. Metabolism results in oxidation of the methyl groups and subsequent conjugation.

Summary

1,2,4 -Trimethylbenzene and other trimethylbenzene isomers are simple aromatic chemicals that do not carry alerts for genotoxic activity.  Trimethylbenzenes have been shown to have no significant genotoxic activity in vitro and in vivo. These observations indicate that trimethylbenzenes are not expected to be genotoxic carcinogens.

There are no data from toxicokinetics, metabolism and distribution findings to indicate any target for possible genotoxic carcinogenic activity.

A number of trimethylbenzenes have been examined in repeat dose inhalation studies and shown low toxicity.  The liver and kidney showed changes consistent with an adaptive change to TMB administration. The modes of action leading to such adaptive changes are such that any tumours resulting on chronic exposure to TMBs would not arise though a genotoxic mechanism and would either be of no relevance to humans (kidney), or of limited relevance (liver), and would be encompassed by the standard toxicological assessment process.

It is concluded that there are sufficient data to indicate that trimethylbenzenes do not pose a risk of genotoxic carcinogenicity.  There are also sufficient data to understand that the cellular changes arising from repeated administration of trimethylbenzenes to animals are minimal and to conclude that trimethylbenzenes are unlikely to pose a significant carcinogenic hazard to humans. The conduct of animal carcinogenicity studies to further evaluate this is considered to be unwarranted.

References

Ashby J and Tennant RW (1988) Chemical structure, Salmonella mutagenicity and extent of carcinogenicity as indicators of genotoxic carcinogenesis among 222 chemicals tested in rodents by the U.S.NCI / NTP. Mutat Res 204: 17-115.

 

Justification for classification or non-classification

1,2,4-Trimethylbenzene has a low potential for carcinogenicity and therefore does not warrant classification under DSD or CLP.