Introduction

1. 1,3-Dichloropropan-2-ol (1,3-DCP) and 2,3 dichloropropan-1-ol (2,3-DCP) are contaminants of some foodstuffs and of polyamine flocculants used in the treatment of drinking water. Both the COC and COM have previously published statements on the closely related compound 3-chloro-1,2-propanediol (3-MCPD)^(1,2) 1,3-DCP and 2,3-DCP were considered by the COC and COM in 2001.^(3,4,) In 2001, the COM recommended that appropriate in-vivo mutagenicity studies should be undertaken with 1,3-DCP and 2,3DCP in accordance with the COM guidelines.^(3,5) In 2001, the COC came to the following conclusions:

It is prudent to assume that 1,3-DCP is a genotoxic carcinogen and that exposures to 1,3-DCP should be reduced to as low a level as technologically feasible.

It is prudent to assume that 2,3-DCP may posses genotoxic activity in-vivo. Although no carcinogenicity data are available, it would however be prudent to reduce exposures to 2,3-DCP to as low a level as technologically feasible.

2. Both of these compounds have been recently considered by the COM who has updated its advice on the mutagenicity of 1,3-DCP and 2,3-DCP in the light of results from new in-vivo mutagenicity studies on these two compounds. An updated COM statement on 1,3-DCP was published in October 2003 and an updated statement on 2,3-DCP was published in June 2004.^(6,7)

3. The available carcinogenicity and other relevant toxicological information on these two compounds including the recent conclusions reached by the COM are considered below. 1,3-DCP and 2,3-DCP have been considered separately in the following sections.

1,3-DCP

4. Available toxicology, mutagenicity and carcinogenicity data for 1,3-DCP has been summarised by the Joint FAO/WHO Committee on Food Additives (JECFA)⁽⁸⁾ although much of the key data remain unpublished. From a 13-week oral toxicity study, a NOAEL of 1mg/kg/day had been identified. Limited information on the metabolism of 1,3-DCP indicates that it may be metabolised to form epichlorohydrin, which may, via glycidol, be conjugated to form mercapturic acid derivatives.⁽⁹⁾ In-vitro investigations with hepatocyte cultures indicate also a pathway involving CYP2E1 to dichloroacetone (a directly acting cytotoxic compound) leading to glutathione depletion.^(10-14)

Mutagenicity

Updated Advice from COM 2003

5. The metabolism of 1,3-DCP was likely to produce a reactive epoxide intermediate that could damage DNA. Members were aware that 1,3-DCP had been found to be mutagenic to Salmonella typhimurium strains TA1535 and or TA 100.^(15-22) Studies with mammalian cells have produced increased frequencies of sister chromatid exchanges and chromosome aberrations. ^(23,24) A positive result has been obtained in a mouse lymphoma assay. ^(25,26) 1,3-DCP was negative in the wing spot test in Drosophila melanogaster (a somatic mutation and recombination test)⁽²⁷⁾.

6. The Committee considered two new in-vivo genotoxicity studies at its May 2003 meeting.^(28,29) These comprised a rat bone-marrow micronucleus test²⁸ and a rat liver unscheduled DNA synthesis (UDS) assay,²⁹ both of which are widely used to assess genotoxicity in vivo.

7. The Committee concluded that both the rat bone-marrow micronucleus test and the rat liver UDS test had been carried out to an acceptable standard and were negative. Thus the additional information recommended by the COM as being necessary to provide adequate reassurance that the mutagenic activity seen in vitro was not expressed in vivo had now been provided. The COM noted the uncertainties with regard to routes of metabolic activation of 1,3-DCP and agreed that the two new mutagenicity studies supported the view that reactive metabolites, if formed, did not produce genotoxicity in vivo in the tissues assessed. The COM concluded that 1,3-DCP can be regarded as having no significant genotoxic potential in vivo.

Carcinogenicity

Advice from COC 2001

8. A 104-week toxicology and carcinogenicity study with 1,3-DCP in Wistar rats was previously considered by COC in 1991.⁽³⁰⁾ At the time COC concluded that 1,3-DCP was genotoxic and carcinogenic, although a formal committee statement was not issued. Additional information on the study was presented to the COC in 2001. The COC concluded that the spectrum of tumours observed in the 104-week rat study (which are reproduced below for ease of access), particularly in the liver and tongue was evidence of a clear carcinogenic effect of 1,3-DCP. It was possible that the tumours in the male kidney could be associated with the high rate of chronic progressive nephropathy observed in the study and additionally, the thyroid follicular cell tumours could be associated with hyperplasia, a toxic finding commonly seen in male rats, although no specific mechanism data were available.

9. A brief summary of the evidence for carcinogenicity of 1,3-DCP in the rat carcinogenicity study provided to COC in 2001 is given below.

• In the liver, combined incidences of hepatocellular adenoma and carcinoma, showed a statistically significant dose-related increase (p<0.001) in both males and females. (eg males -controls 1/50, high dose 8/50; females - controls 1/50, high dose 41/50).
• In the tongue, combined incidences of squamous cell papilloma and carcinoma showed a statistically significant dose-related increase (p<0.001) in both males and females. (eg males - controls 0/50, high dose 12/50; females - controls 0/49, high dose 11/49).
• In the thyroid combined incidences of follicular cell adenoma and carcinoma showed a statistically significant dose-related increase (p<0.001) in both males and females (eg males - controls 0/50, high dose 4/50; females - controls 1/49, high dose 5/49).
• In the kidney, combined incidences of renal tubular adenoma and carcinoma, showed a statistically significant dose-related increase (p<0.001) in males only (eg controls 0/50, high dose 9/50).
  

Updated Advice from COC 2004

10. The COC reaffirmed its previous opinion that 1,3-DCP induced tumours of the kidney and thyroid could have been secondary to sustained cell proliferation. Members also agreed that there was evidence of a hepatotoxic effect at doses below those producing a significant increase in combined hepatocellular adenoma and carcinoma. The Committee agreed that the evidence of hepatotoxicity, together with negative results from the in vivo rat liver UDS assay, provided evidence of non-genotoxic mode of action in the liver.

Further consideration of tumours of the tongue (November 2003 and April 2004 meetings)

11. The Committee then considered possible modes of action of 1,3-DCP in inducing tumours of the tongue. The Committee considered that the finding of 2% papillary carcinoma in the mid-dose female group might not be treatment related but the incidence of tongue papilloma (14.3%) and carcinoma (8.2%) in high dose females was clearly treatment related. In male rats the incidence of tongue tumours in the high dose group was 12% (for both papilloma and carcinoma). There were no tongue tumours in males at the low and mid dose groups. The high dose level clearly exceeded the Maximum Tolerated Dose level in that there was an increase in treatment related mortality and hepatotoxicity. The Committee agreed that 1,3-DCP was an irritant and had produced irritant effects in gastric mucosa of treated rats, but there were no suitable data on the potential for 1,3-DCP irritation of the tongue. Members noted that at the time of conduct of the bioassay (1986) it was not routine to examine the tongue histologically. It was agreed however, that since the compound had been given in the drinking water in the bioassay, chronic irritation was a plausible hypothesis for the induction of the tumours in the tongue. Members discussed the suggestion that bacteria metabolised 1,3-DCP to the genotoxic carcinogen epichlorohydrin but agreed there was no specific evidence to support this proposal.

12. The COC discussed future research to investigate whether the tumours of the tongue occurred via a genotoxic mechanism and agreed that information on contact-irritancy, cell proliferation and formation of DNA adducts in tongue tissue using ³²P-postlabelling in animals treated with suitably high doses of 1,3-DCP was needed.

13. The COC concluded that until such data were available, it was not possible to exclude the possibility of a genotoxic mechanism for the tumours of rat tongue seen in a long-term drinking water study with 1,3-DCP.

2,3-DCP

14. There are very little data on the absorption, distribution, and excretion of 2,3-DCP. Theoretically, 2,3-DCP could be metabolised to produce epichlorohydrin (and subsequently glycidol) and therefore there were structural alerts for genotoxicity and carcinogenicity. One research group had provided some in vitro data to suggest that induction of CYP2E1 resulted in 2,3-DCP mediated hepatotoxicity and glutathione depletion.⁽³¹⁾ The findings of Koga et al⁽³¹⁾ suggest dechlorination/ hydroxylation of 2,3-DCP may occur but the evidence for epoxide formation was not conclusive. There are insufficient data to draw conclusions on the metabolic activation of 2,3-DCP but overall the evidence suggested metabolic activation of 2,3-DCP differs from 1,3-DCP. The COM considered these data and agreed that that the metabolism of 2,3-DCP had not been fully elucidated. Metabolic activation in vivo to active metabolites had been postulated but had not been proven.

Mutagenicity

15. The COM concluded in 2001 that 2,3-DCP was mutagenic in vitro in Salmonella typhimurium strains TA 100 and TA 1535 in a study with and without metabolic activation⁽¹⁸⁾ and mutagenic in another Ames test.⁽¹⁹⁾ Positive results were also obtained for sister chromatid exchange with Chinese Hamster V79 cells both with and without metabolic activation.⁽²³⁾

Updated advice from COM 2004

16. The COM considered two new in-vivo genotoxicity studies at its February 2004 meeting. These comprised a rat bone-marrow micronucleus test⁽³²⁾ and a rat liver unscheduled DNA synthesis (UDS) assay,⁽³³⁾ both of which are widely used to assess genotoxicity in vivo.

17. The Committee concluded that both the rat bone-marrow micronucleus test and the rat liver UDS test had been carried out to an acceptable standard and were negative. Thus the additional information recommended by the COM as being necessary to provide adequate reassurance that the mutagenic activity seen in vitro was not expressed in vivo had now been provided. The Committee noted the uncertainties with regard to routes of metabolic activation of 2,3-DCP and agreed that the two new mutagenicity studies supported the view that reactive metabolites, if formed, did not produce genotoxicity in vivo in the tissues assessed The Committee concluded that 2,3-DCP can be regarded as having no significant genotoxic potential in vivo.

Carcinogenicity

18. Further advice on the carcinogenicity of 2,3-DCP was sought from the COC at the June 2004 meeting in the light of the updated advice on mutagenicity from COM.

19. Although there are no carcinogenicity studies available for 2,3-DCP, the World Health Organisation's International Agency for Research on Cancer (IARC) evaluated the brominated analogue, 2,3 dibromo-propanol (2,3 DBP) and considered that "there is sufficient evidence in experimental animals for the carcinogenicity of 2,3 dibromo propan-1-ol". In addition skin application of 2,3 DBP produced multisite tumours in both rats and mice.³⁴ However, the Committee considered this information in 2001 and agreed that no conclusions could be drawn from the studies on 2,3 dibromo propan-1-ol in respect of the carcinogenicity of 2,3-DCP.

20. Thus no conclusions regarding the carcinogenicity of 2,3-DCP can be reached on the available information on this compound.

Conclusions

21. The Committee concluded:

1,3-DCP: The COC concurs with its previous advice that 1,3-DCP should be regarded as a genotoxic carcinogen. It is not possible to exclude a genotoxic mechanism for the induction of the tumours of rat tongue seen in a long-term drinking water study with 1,3-DCP. The Committee recommended that further investigations regarding the mechanism of 1,3-DCP carcinogenicity in the rat tongue should include information on contact-irritancy, cell proliferation and formation of adducts in tongue tissue using ³²P-postlabelling in animals treated with suitably high doses of 1,3-DCP.

2,3-DCP: The available evidence is consistent with the conclusion that 2,3-DCP does not posses genotoxic activity in-vivo. There are no appropriate carcinogenicity bioassays of 2,3-DCP available. No conclusions regarding carcinogenicity of 2,3-DCP can be reached.

June 2004

References

1. COM statement on 3-monochloro propane 1, 2-diol (3-MCPD) (Revised October 2000) http://www.advisorybodies.doh.gov.uk/com/index.htm

2. COC (2000). Statement on carcinogenicity of 3-monochloro propane 1,2- diol (3-MCPD) COC statement COC/00/S5 - December 2000
(update of COC/99/S5) http://www.advisorybodies.doh.gov.uk/mcpd1.htm

3. COM (2001). Statement on 1,3-dichloropropan-2-ol and 2,3 dchlororpropan-1-ol. COM Statement COM/01/S2.

4. COC (2001). Statement on Carcinogenicity of 1,3-dichloropropan-2-ol (1,3 DCP) and 2,3-dichloropropan-1-ol (2,3 DCP) COC statement COC/01/S1 - May 2001

5. COM (2000). Guidance on a Stategy for the Testing of Chemcials for Mutagenicity. http://www.advisorybodies.doh.gov.uk/com/guidance.pdf

6. COM (2003). Statement On The Mutagenicity Of 1,3-Dichloropropan-2-ol Com/03/S4 - October 2003.

7. COM (2004). Statement on the mutagenicity of 2,3-Dichloropopan-1-ol. COM/04/S1.

8. Olsen P. (1993) Chloropropanols In: Toxicological Evaluation of Certain Food Additives and Contaminants, Joint Expert Committee on Food Additives, World Health Organization, Geneva, Switzerland, (41st Meeting) (WHO FOOD ADDITIVES SERIES) No. 32:267- 285.

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14. Fry JR, Sinclair D, Holly Piper C, Townsend S-L, Thomas NW. (1999) Depression of glutathione content, elevation of CYP2El -dependent activation, and the principal determinant of the fasting-mediated enhancement of 1,3-dichloro-2-propanol hepatotoxicity in the rat. Food Chem Toxicol 37;351-355.

15. Hahn H. Eder E and Deininger C. (1991). Genotoxicity of 1,3-dichloro-2-propanol in the SOS chromotests and in the Ames tests. Elucidation of the genotoxic mechanism. Chem. Biol. Interactions, 80: 73-88.

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17. Stolzenberg S.J. & Hine C.H. (1980) Mutagenicity of 2- and 3-carbon halogenated compounds in the Salmonella/mammalian-microsome test. Environmental Mutagenesis, 2: 59-66.

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20. Ohkubo T, Hayashi T, Watanabe E, Endo H, Goto S, Mizoguchi T, Mori Y (1995) Mutagenicity of chlorohydrins. Nippon Suisan Gakkaishi 61: 596-601 (in Japanese).

21. Gold MD, Blum A and Ames B (1978). Another flame retardant, Tris-(1,3-dichloro-2-propyl)-phosphate, and its expected metabolites are mutagens. Science 200: 785-787.

22. Lynn RK, Wong K Garvie-Gould C and Kennish JM. (1981) Disposition of the flame retardant, Tris (1,3-dichloro-2-propyl) phosphate in the rat. Drug metabolism and Disposition, 9: 434-451.

23. von der Hude W. Scheutwinkel M, Gramlich U, Fissler B and Basler A (1987). Genotoxicity of three-carbon compounds evaluated in the SCE test In-Vitro. Environmental Mutagenesis, 9: 401-410.

24. Putman D. & Morris M. (1990). Sister chromatid exchange and chromosome aberration assay in Chinese hamster ovary cells, 1,3-dichloro-2-propanol. Microbiological Associates, Inc. Laboratory study No. T9250.337 NTP.

25. San R. & Blanchard M. (1990). L5178Y TK^+/- mouse lymphoma mutagenesis assay, 1,3-dichloro-2-propanol. Microbiological Associates, Inc. Laboratory study No. T9250.702.

26. Henderson LM, Bosworth HJ, Ransome SJ, Banks SJ, Brabbs CE and Tinner AJ . (1987) An assessment of the mutagenic potential of 1,3-dichloro-2-propanol, 3-chloro-1,2-propanediol and a cocktail of chloropropanols using the mouse lymohoma TK locus assay. Unpublished report No ULR 130 ABC/861423 from Huntingdon Research Centre, Huntingdon, Cambridgeshire England.

27. Frei H. & Wurgler F (1997) The vicinal chloroalcohols 1,3-dichloro-2-propanol (DC2P), 3-chloro-1,2-propanediol (3CPD) and 2-chloro-1,3-propanediol (2CPD) are not genotoxic in-vivo in the wing spot test of Drosophila melanogaster. Mut Res 394: 59-68. 16.

28. Howe, J. (2002) 1,3-Dichloropropan-2-ol (1,3-DCP): Induction of micronuclei in the bone marrow of treated rats. Report no 2150/1-D6172 from Covance Laboratories Ltd, Harrogate, North Yorkshire, England. Available from the Food Standards Agency.

29. Beevers, C. (2003) 1,3-Dichloropropan-2-ol (1,3-DCP): Induction of unscheduled DNA synthesis in rat liver using an in vivo/in vitro procedure. Report no 2150/3-D6173 from Covance Laboratories Ltd, Harrogate, North Yorkshire, England. Available from the Food Standards Agency.

30. Hercules Inc. (1986) 104-Week Chronic Toxicity and Oncogenicity Study with 1,3-Dichloropropan-2-ol in the Rat. Unpublished Report No. 017820 from Research and Consulting Company AG, ltingen, Switzerland.

31. Koga M, Inoue N, Imazu K, Yamada N and Shinoki T (1992). Identification and quantitative analysis of urinary metabolites of dichloropropanols in rats. J UOEH (University Occupational and Environmental Health, 14, 13-22.

32. Beevers C. (2003) 2,3-Dichloropropan-2-ol (1,3-DCP): Induction of micronuclei in the bone marrow of treated rats. Report no 2150/2-D6172 from Covance Laboratories Ltd, Harrogate, North Yorkshire, England. Available from the Food Standards Agency.

33. Beevers, C. (2003) 2,3-Dichloropropan-2-ol (1,3-DCP): Induction of unscheduled DNA synthesis in rat liver using an in vivo/in vitro procedure. Report no 2150/4-D6172 from Covance Laboratories Ltd, Harrogate, North Yorkshire, England. Available from the Food Standards Agency.

34. IARC (2000). IARC Monographs on the evaluation of carcinogenic risks to humans .2,3 Dibromopropan-1-ol Vol 77 p439-454.

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