Furfuryl alcohol

Endpoint summary

Additional information

The genotoxic potential of furfuryl alcohol was reviewed and reported in an EFSA review (2011). A Comet Assay has been conducted and some additional relevant data have been found in an updated literature review and are included herein along with the key studies from the EFSA review.

Non-human information

In vitro data

The key studies are considered to be bacterial mutation assays (Mortelmans et al, 1986, Aeschbacher et al, 1989) and mammalian cell cytogenetic assays (Galloway et al, 1987, Stich et al, 1991). These are two recognised core assay types for investigating mutagenicity in vitro.

Furfuryl alcohol was tested using a pre-incubation Ames assay in S. typhimurium strains TA1535, TA1537, TA100, and TA98 in both the presence and absence of rat and hamster liver auxiliary metabolic activation (S9). A range of doses up to 10000 ug/plate was used. Furfuryl alcohol was negative in this assay (Mortelmans et al, 1986). Aeschbacher et al (1989) extended the range of strains used to include TA102. This study used a pre-incubation Ames assay with S. typhimurium strains TA98, TA100 and TA102 in both the presence and absence of rat liver S9. A range of doses up to 1 mmol/plate was used. Furfuryl alcohol was again negative in this assay. Additional studies reporting Ames assay results are included in an EFSA review (EFSA 2011), indicating two negative studies and one positive study. However, the positive result is reported at a dose of 200000 ug/ml. A recent study by Monien (2011) has reported a positive response in a Salmonella strain TA100 modified to express human sulphotransferase 1A1. The weight of evidence is that furfuryl alcohol is not genotoxic in standard bacterial mutation assays, but is genotoxic in a strain with increased sulphotransferase activity.

In a mammalian cell cytogenetic assay (Galloway et al 1987), furfuryl alcohol was examined in cultured CHO cells at a range of doses up to 500 ug/mL in the absence of rat liver S9 and 1600 ug/mL in the presence of S9. Cells were arrested in metaphase 2 hours following a 10 hour exposure -S9, and 11 hours following a 2 hour exposure +S9, and evaluated for chromosomal damage. Furfuryl alcohol was negative in the absence of S9. In the presence of S9, a small increase in chromosomal aberrations was noted, but this was not reproduced in a repeat experiment, which also included higher dose levels. The report concluded an equivocal result. Stich et al (1981) also examined furfuryl alcohol in CHO cells in both the absence and presence of rat liver S9. Exposure was for 3 hours (both + and -S9) and sampling was after a further 20 hour culture. In the absence of S9, an increase in aberrations was seen only at dose levels of 10 mM or above, whereas in the presence of S9, increases were seen at concentrations of 2.5 mM.

A negative result for chromosomal aberration induction was reported by Sujatha (2008) following treatment of human peripheral lymphocytes in the absence of S9. However, the doses selected for this study were not based on maximum tolerated doses from an in vitro preliminary test, but were calculated as relevant to (corresponding to) an in vivo study previously performed.

Additional data for furfuryl alcohol from non-core genotoxicity assays in vitro are summarised by an EFSA review (EFSA, 2011) and include an examination for the ability to induce sister chromatid exchanges (SCE), with a reported positive result in CHO cells, but negative results in two studies with human lymphocytes. However, a further positive result for SCE induction in human lymphocytes was reported by Sujatha (2008). Positive reports are also given for gene conversion in yeast and DNA repair in bacteria. These non-core studies are considered to be of limited value in a weight of evidence evaluation compared with the core genotoxicity assays above. An investigative study has reported DNA adducts in bacterial DNA following exposure of a Salmonella strain modified to have enhanced expression of suphotransferase activity (Monien, 2011).

The findings from the standard in vitro genotoxicity reports indicate that furfuryl alcohol is negative in gene mutation assays (Ames test), but shows some activity in cytogenetic assays. These effects appear to be of limited reproducibility. Furfuryl alcohol is genotoxic in a Salmonella strain modified to express enhanced levels of sulphotransferase activity.

 

In vivo data

The key studies are considered to be cytogenetic studies in the mouse examining for bone marrow micronuclei (NTP 1999) and chromosomal aberrations (NTP, 1999) and a Comet assay examining DNA strand breaks and cytotoxicity in the glandular stomach, liver and kidney of male CD1 mice (Barfield, 2016). These are recognised core assay types for investigating mutation in vivo.

For the micronucleus assay, furfuryl alcohol was administered intraperitoneally to mice (5 per group) at a range of doses up to 215 mg/kg on three successive days and the bone marrow examined 24 hours after the last dose. Furfuryl alcohol gave a negative response. For the chromosomal aberration assay furfuryl alcohol was administered by a single intraperitoneal injection to mice (8 per group) at a range of doses up to 300 mg/kg and the bone marrow examined at 17 hours and 36 hours after dosing. Furfuryl alcohol gave a negative response. The Comet assay used doses of 93.8, 187.5 and 375 mg/kg/day dosed orally on two consecutive days. Animal were killed 3 hours after the second dose and cell suspensions from glandular stomach, liver and kidneys prepared and electrophoresed. There were no increases in the median % tail intensity compared to vehicle control and no hedgehog "ghost" cells. Furfuryl alcohol was negative in this Comet assay for all three tissues sampled.

In an earlier study, Sujatha and Subramanyam (1994) reported examining furfuryl alcohol by oral gavage to mice either as a single dose or as five daily doses of 1000, 2000 and 4000 ppm in water, with sampling at various times later. They reported statistically significant increases in chromosomal aberrations (excluding gaps) at the highest dose tested at 18, 24 and 36 hour sampling times after a single dose and at 12, 18, 24 and 36 hour sampling time after repeated dosing. There was also a statistically significant increase reported after a single dose of 2000 ppm at the 18 hour sampling time. This study used only 2 animals per group. Sujatha (2007) used a similar protocol to examine for SCE induction in mice and reported small (approximately 2x control maximum) increases over controls. The study again used only 2 animals per group.

Additional data for furfuryl alcohol from non-core genotoxicity assays in vivo are summarised by an EFSA review (EFSA 2011) and include negative results for SCE induction in mice, and for mutation in drosophila. A recent study by Monien (2011) reported low levels of DNA adducts formed in liver, kidney and lung tissue, but not colonic mucosa from mice exposed to furfuryl alcohol for 28 days in the drinking water at approximately 390 mg/kg.

Human information

An examination of lymphocytes from adults in an occupational exposure setting reported a negative result for SCE induction (Gomez-Arroyo & Souza, 1985). Exposure levels of 32300 mg/m3 to 97120 mg/m3 furfuryl alcohol were reported. The workers on the plant were exposed to both furfuryl alcohol and to furfural.

 

Summary and Discussion of Mutagenicity

Furfuryl alcohol has been examined for mutagenicity both in vitro and in vivo in a range of recognised core assay types, and also in a number of non-standard genotoxicity assays of limited relevance for a weight of evidence assessment of genotoxic activity.

In vitro, furfuryl alcohol has been examined for gene mutation in a number of studies using the Ames test. It can be concluded to be non-genotoxic for this endpoint in standard tester strains, but genotoxic in a Salmonella strain modified to have enhanced sulphotransferase activity. DNA adducts were found in the DNA from such modified bacteria. There is conflicting evidence for clastogenic activity from in vitro mammalian cell assays, but with some positive results recorded. These have been of limited reproducibility however, and the effects seen were sometimes small. Overall, there is some limited evidence for cytogenetic activity in vitro.

In vivo, there is evidence from a robust evaluation in mice showing negative results in the bone marrow for the endpoints of micronucleus, chromosomal aberration and SCE induction and from a Comet assay. A report of a positive result for chromosomal aberrations was from a study using two animals per group, and is considered insufficient to alter a conclusion of no clastogenic activity in vivo. A recent study by Monien (2011) reported DNA adducts formed in liver, kidney and lung tissue, but not colonic mucosa from mice exposed for 28 days to furfuryl alcohol.

Other publications (Hoie, 2015; Sachse et al, 2014, 2016a, b) using different routes of administration and different modifications of sulphotransferase activity also report of finding of adducts in mice. These data do not affect the overall interpretation.

It is concluded that the available data indicate that furfuryl alcohol has no significant mutagenic activity in the conventional endpoint assays examined. Modification of Salmonella to artificially enhance sulphotransferase activity, however, resulted in DNA adducts and a mutagenic response in vitro. Investigation of mice dosed with furfuryl alcohol resulted in the same DNA adducts being detected in some but not all of the tissues examined, and only at low levels. The significance, if any, of this DNA binding (an indicator assay for genotoxicity) is not clear, although furfuryl alcohol was not mutagenic in two endpoint studies for mutagenicity in the bone marrow, and was also negative in the comet assay in three tissues, including two where adducts had previously been reported.

It is concluded that there is limited evidence for genotoxicity of furfuryl alcohol in vitro, but no evidence for mutagenicity in vivo.

Additional references

EFSA. EFSA Journal 2011; 9(3):1840

Galloway SM, Armstrong MJ, Reuben C, Colman S, Brown B, Cannon C, Bloom AD, Nakamura F, Ahmed M, Duk S, Rimpo J, Margolin BH, Resnick MA, Anderson B and Zeiger E (1987). Chromosome aberrations and sister chromatid exchanges in Chinese hamster ovary cells: Evaluations of 108 chemicals. Environ. Mol. Mutagen. 10 (Suppl. 10), 1-175.

Høie AH, Monien BH, Sakhi AK, Glatt H, Hjertholm H, Husøy T (2015). Formation of DNA adducts in wild type and transgenic mice expressing human sulfotransferases 1A1 and 1A2 after oral exposure to furfuryl alcohol. Mutagenesis 30 (5): 643-9.

Sachse B, Meinl W, Glatt H, Monien BH (2014).The effect of knockout of sulfotransferases 1a1 and 1d1 and of transgenic human sulfotransferases 1A1/1A2 on the formation of DNA adducts from furfuryl alcohol in mouse models. Carcinogenesis 35 (10): 2339-2345.

Sachse B, Meinl W, Sommer Y, Glatt H, Seidel A, Monien BH (2016a).Bioactivation of food genotoxicants 5-hydroxymethylfurfural and furfuryl alcohol by sulfotransferases from human, mouse and rat: a comparative study. Archives of Toxicology 90 (1), 137-148.

Sache B, Meinl W, Glatt H, Monien BH (2016b). Ethanol and 4-methylpyrazole increase DNA adduct formation of furfuryl alcohol in FVB/N wild-type mice and in mice expressing human sulfotransferases 1A1/1A2. Carcinogenesis 37 (3): 314-9.

Short description of key information:
Furfuryl alcohol has been examined for mutagenicity both in vitro and in vivo in a range of recognised core assay types, and also in a number of non-standard genotoxicity assays of limited relevance for a weight of evidence assessment of genotoxic activity.
It is concluded that there is limited evidence for genotoxicity of furfuryl alcohol in vitro, but no evidence for mutagenicity in vivo, from the studies to date.

Endpoint Conclusion: No adverse effect observed (negative)

Justification for classification or non-classification

It is concluded that the available data are sufficient to indicate that furfuryl alcohol has no significant genotoxic activity and does not warrant classification under DSD or CLP.

This conclusion is endorsed by the position reached by the Technical Committee on C&L of dangerous substances (European Chemicals Bureau, ECBI/90/06 Rev. 8) which agreed not to classify furfuryl alcohol for genotoxicity. The EFSA (2011) also concludes on a lack of concern for genotoxicity.