4.3.2.3. 1,4-dioxane

*Occurrence: 1,4-dioxane* (1,4-diethylene dioxide) is an oxygen-containing single ring molecule (Figure 8(5)), which is mainly used as a solvent and stabilizer [558]. It can occur in some foods, including meat, tomatoes, shrimp and coffee as a natural constituent, as a contact material from food packaging or contaminated water, or as an impurity in food additives, such as polyethylene glycol and polysorbate [815–817]. Analysis of food products in Japan, revealed that content of 1,4-dioxane ranged from 3 to 13 μg/kg [818].

*Carcinogenicity:* Administration of 1,4-dioxane in drinking water induced hepatocellular adenomas or carcinomas in rats and mice of both sexes and in male guinea pigs [68,558,815,816,819,820]. Other target organs of carcinogenicity included nasal cavity in rats of both sexes, mammary gland in female and abdominal cavity in male rats, gallbladder in male guinea pigs. Administration of 1,4-dioxane in drinking water at 25,000 ppm to rats of both sexes in a 13-week study induced glutathione S-transferase (GST) placental form-positive hepatocellular foci, which are known preneoplastic lesions [821]. In addition, 1,4-dioxane promoted hepatocellular foci produced by administration of diethylnitrosamine [822].

*Genotoxicity/DNA Binding (Adducts):* 1,4-dioxane was non-genotoxic in vitro, in vivo genotoxicity studies in rodents were also mainly negative although some positive results suggesting weak genotoxicity were observed at high cytotoxic doses exceeding 1500 mg/kg [815,816,823–826]. More recent studies provide evidence that 1,4-dioxane induced chromosomal breaks, DNA damage and mutagenicity in the liver of rats or mice [827–830]; however, these studies also used high dose levels of 1,4-dioxane (above 1000 mg/kg) and thus, no clear conclusion concerning genotoxicity of 1,4-dioxane can be made. DNA adductome analysis detected several DNA adducts with unidentified chemical structure after administration of 1,4-dioxane to male rats in the drinking water at 200 and 5000 ppm; however, these adducts could have resulted from oxidative damage, rather than direct covalent binding [831].

*Metabolism:* 1,4-dioxane is mainly metabolized in the presence of mixed-function oxidases to 1,4-dioxane-2-one and then to β-hydroxyethoxyacetic acid, which is excreted in urine in rats and humans [558,815,816,832]. Induction of CYP2B1/2, CYP2C11, and CYP2E1 in the liver and only CYP2E1 in the kidney and nasal mucosa was observed in rats exposed to 1,4-dioxane in the drinking water, while dosing by gavage induced CYP3A activity in the liver [833].

*MoA:* The mechanism(s) of carcinogenicity of 1,4-dioxane has not been elucidated but is unlikely to involve genotoxicity. Studies suggested that hepatocarcinogenicity of 1,4 dioxane likely results from cytotoxicity followed by regenerative hyperplasia, in addition, mitogenic response was suggested as a key initiating event [815,834–837]. Such effects are threshold-dependent. Oxidative damage might also play a role [838]. Tumors of nasal passages were attributed to inhalation of drinking water containing 1,4-dioxane [815,839]. SCF [817] concluded that since 1,4-dioxane is likely to exert its carcinogenic effects by non-genotoxic mechanisms, use of a threshold approach to determine acceptable levels of exposure is justified.

*Human Exposure:* Dietary exposures to 1,4-dioxane is a minor exposure route, in contrast to inhalation. FDA [840] estimated per capita dietary intake of 1,4-dioxane to be low, averaging at 0.6 μg/person/day. Analyses of Japanese foods revealed an EDI of 1,4-dioxane averaging from 0.44 to 4.5 μg/kg bw/day [818,841]. SCF established that an estimated maximum exposure to 1,4-dioxane as a constituent of food additives, polysorbates, in bread ranges between 0.008 to 0.05 μg/kg bw/day [817].

*Human Effects:* No epidemiological studies investigating association of oral exposure of humans to 1,4-dioxane and cancer are currently available [68,558,816]. Limited occupational studies found no excess of death from cancer associated with 1,4-dioxane exposure [815].

*Risk:* IARC [558] assigned 1,4-dioxane to a group of chemicals which are possibly carcinogenic to human (Group 2B) (Table 2). SCF [817] established that exposure to 1,4 dioxane in food additives is significantly lower than the established NOAEL of 10 mg/kg bw/day, and thus is of no toxicological concern.

## 4.3.2.4. Methyl Isobutyl Ketone

*Occurrence*: *Methyl isobutyl ketone* (MIBK) (4-methylpentan-2-one) (Figure 8(5)) is produced from acetone by aldol condensation and is used primarily as denaturant and solvent [115,842,843]. It is also a natural constituent of many foods including orange and lemon juice, grapes, papaya, ginger, cooked eggs, meat, milk and cheeses, beer, mushrooms, coffee and tea, as at concentrations ranging from 0.008 to 6.5 mg/kg, and as a flavoring agent in meat products, dairy and non-alcoholic beverages, baked goods and puddings at maximum reported level of 25 mg/kg [115,844]. As a component of adhesive, MIBK can also migrate into foods from packaging at levels around 10 to 12 mg/kg.

*Carcinogenicity*: Currently, no studies assessed carcinogenicity of MIBK following oral exposure. Some evidence of carcinogenic activity were observed in inhalation studies, which reported increased incidences of renal tubule neoplasms in rats and hepatocellular neoplasms in mice at the highest tested dose of 1800 ppm (equivalent to 1725 mg/kg/day for rats and 3171 mg/kg/day for mice) [115,842,844–846].

*Genotoxicity/DNA Binding (Adducts)*: MIBK produced overwhelmingly negative results in genotoxicity testing battery in vitro and in vivo [115,843,846–849] and thus, is not considered to be of concern for genotoxicity [842,844].

*Metabolism*: MIBK is metabolized in vivo by reduction in the presence of alcohol dehydrogenases to 4-hydroxyMIBK and by oxidation in the presence of CYP-dependent monooxygenase to 4-methyl-2-pentanol [843,850–853]. The letter metabolite was not detected with oral administration [852,854]. MIBK has been shown to induce liver and renal CYPs, potentiating hepato- and nephrotoxicity produced by chloroform and carbon tetrachloride [853,855,856].

*MoA*: In the carcinogenicity bioassay, histopathologic changes observed in the kidneys of rats were characteristic of α2u-globulin nephropathy [846], suggesting that α2u-globulinmediated MoA is involved in renal carcinogenesis [275,845,857,858]. This MoA is not considered relevant to humans [73]. MoA underlying hepatocarcinogenicity of MIBK in mice is not well understood. While it potentiated hepatotoxicity and cholestasis produced by other chemicals [853,855,856,859,860] no evidence of hepatotoxicity was observed when

MIBK was administered alone [275,843]. A study by Hughes and colleagues [861] suggested involvement of receptor-mediated mechanism, specifically, activation of the CAR/PXR nuclear receptors, which results in hepatocellular proliferation consequently leading to tumor development.

*Human Exposure*: Dietary per capita exposure toMIBK was calculated to be 7 μg/person/day in Europe and 2 μg/person/day in USA [849]. More recent estimations suggest lower levels of intake of 0.02 μg/kg/day [115].

*Human Effects*: No data on human carcinogenicity of MIBK are currently available [115]. Long term exposure in occupational settings was reported to cause cognitive impairment [862].

*Risk*: IARC [275] classified MIBK as possibly carcinogenic to humans (Group 2B) (Table 2). JECFA [849] concluded that at the current levels of intake as a flavoring agent MIBK is unlikely to pose any hazard to human health.

#### *4.4. Carcinogens Formed during Processing, Packaging and Storage of Food*

Food processing contaminants are generated through cooking practices or as a result of food packaging and storage. Some of the carcinogens belonging to this type were discussed in the section on DNA-reactive carcinogens. Examples of epigenetic processing carcinogens in food include alkylated imidazoles and furan.

**Figure 9.** Chemical structures of non-DNA-reactive carcinogens formed during processing, packaging and storage of food.
