First draft prepared by Dr P.J. Abbott
Australia–New Zealand Food Authority, Canberra, Australia
Furfural was evaluated previously by the Committee at its thirty-ninth and fifty-first meetings (Annex 1, references 101 and 137). An ADI was not established at either meeting because of concern about the finding of tumours in male mice given furfural in corn oil by gavage and the fact that no NOEL was identified for hepatoxicity in male rats. In a study in mice, the combined incidence of adenomas and carcinomas was increased in males at the highest dose (175 mg/kg bw per day). In order to address its concern with regard to the formation of liver tumours in mice, the Committee at its fifty-first meeting requested the results of studies of DNA binding or adduct formation in vivo to clarify whether furfural interacts with DNA in the liver of mice, and also requested the results of a 90-day toxicity study in rats to identify a NOEL for hepatotoxicity (Annex 1, reference 137).
Since the last meeting, the results of a 14-day study to determine a dose range, a 90-day study of toxicity in rats, and an assay for unscheduled DNA synthesis in mice in vivo have become available. These data were reviewed and are summarized in the following monograph addendum.
In a 14-day range-finding study with microencapsulated furfural in a carrier of maltodextrin and mixed sugars, groups of five male and five female Fischer 344/N rats were fed a diet containing furfural at concentrations providing doses of 0, 30, 60, 90, 120, and 180 mg/kg bw per day. An additional control group received basal diet containing the encapsulation material only. The animals were examined daily, and body weights and food consumption were recorded weekly. Necropsy was performed at 14 days, and the tissues were examined histologically. Clinical chemical and urinary parameters were examined.
There were no clinical signs of toxicity, and the body weights and food consumption were normal in all groups. Cholesterol and phospholipid concentrations were slightly increased in males at the two higher doses, but these changes were not dose-related. Similarly, females at some doses had decreased blood urea nitrogen and creatinine concentrations, but these changes were not dose-related. A significant decrease in the plasma activity of alanine aminotransferase was found in females at the high dose, which corresponded to significant increases in the absolute (111%) and relative (115%) weights of the liver in these animals. The NOEL was 120 mg/kg bw per day (Jonker, 2000a).
In a 13-week study of toxicity, groups of 10 Fischer 344 rats of each sex were fed diets containing microencapsulated furfural (carrier, maltodextrin and mixed sugars) providing a nominal dose of 0, 30, 60, 90, or 180 mg/kg bw per day. The actual doses found by analysis of the food were 0, 26, 53, 82, and 160 mg/kg bw per day for males and 0, 28, 57, 86, and 170 mg/kg bw per day for females (Jonker, 2000b). An additional control group received a diet containing the encapsulated material without furfural. The animals were examined daily for clinical signs of toxicity, and body weights and food consumption were measured weekly. Animals at the high dose and controls underwent an ophthalmoscopic examination, while all animals were examined for clinical parameters. Gross examinations were performed at autopsy, with measurements of organ weights and extensive histological examination of a range of organs. In a brief report, Buck (2000) demonstrated that furfural is rapidly released from the microencapsules in an aqueous environment.
There were no clinical signs of toxicity, and body weights and food consumption were unaffected by treatment. The animals given the high dose showed no ophthalmoscopic changes when compared with controls. Some changes in clinical chemistry were seen. The haematological changes included a decreased red blood cell count in males at 180 mg/kg bw per day and increased corpuscular volume and mean corpuscular haemaglobin in males at 90 and 180 mg/kg bw per day. Females at the high dose showed decreased alkaline phosphatase activity, increased gamma-glutamyltransferase activity, an increased plasma concentration of albumin, and a decreased plasma concentration of potassium. Males at the high dose showed decreased alanine aminotransferase activity, an increased plasma concentration of albumin, and an increased albumin:globulin ratio. Increased albumin:globulin ratios were also found in males at 30 and 90 mg/kg bw per day but not in those at 60 mg/kg bw per day.
At necropsy, the absolute and relative weights of the liver were increased in males at 180 mg/kg bw per day, but there were no gross pathological changes. Microscopic examination revealed changes in the liver in 5/10 males at 90 mg/kg bw per day and in 10/10 at 180 mg/kg bw per day. The changes were found mainly in the perilobular region and were characterized by cells with less coarse cytoplasm, an increased occurrence of clumps of eosinophils, a less dense periphery, and more prominent nucleoli in the nucleus. The changes seen at 90 mg/kg bw per day were not severe, and those in rats at 180 mg/kg bw per day were slight; none were accompanied by signs of degeneration or necrosis. No changes were observed in the livers of females, and there were no signs of hepatotoxicity such as degeneration, necrosis, or inflammation. No bile-duct proliferation was seen. The NOEL was 60 mg/kg bw per day (Jonker, 2000c).
The ability of furfural to induce DNA repair in the hepatocytes of B6C3F1 mice was assessed in an assay for unscheduled DNA synthesis. The maximum tolerated dose for animals of each sex was determined in a preliminary study to be 320 mg/kg bw. In the study of unscheduled DNA synthesis, doses of 50, 175, and 320 mg/kg bw were given to groups of three animals of each sex, and expression of DNA repair was measured 2–4 and 12–16 h after treatment. N-Nitrosodimethylamine (20 mg/kg bw) was used to measure expression within 2–4 h and aminoazotoluene (200 mg/kg bw) for expression within 12–16 h, as positive controls.
The animals treated with furfural did not show increased unscheduled DNA synthesis at either time after dosing, whereas the positive controls showed statistically significant increases in net nuclear grain counts. Little replicative DNA synthesis (0–0.4%) was seen at either interval. The results provided no evidence that furfural damages DNA in mouse hepatocytes at doses up to 320 mg/kg bw (Edwards, 1999).
Dietary intake of furfural from its use as a flavouring agent was estimated from ‘poundage’ data provided by the flavouring industry. The method used for this estimation is described elsewhere in this volume (see monograph on furfural alcohol and related flavouring agents). With this method, the estimated daily intake was 9 µg/kg bw in Europe and 8 µg/kg bw in the USA.
The Committee reviewed the results of a 13-week study in rats in which microencapsulated furfural was administered in the diet. It noted that, in a complementary study, furfural was rapidly and completely released from microencapsulation in an aqueous environment and therefore considered that this form of administration of furfural was suitable for a feeding study. In the 13-week dietary study, minor hepatocellular alterations were observed in males, but not in females, at doses of 82 and 160 mg/kg bw per day. While these changes might be judged not to be adverse, the Committee took a conservative position, and considered the NOEL to be 53 mg/kg bw per day, at which dose there was no evidence of hepatic alterations. This result contrasts with those of the previous studies in rats, in which furfural was administered in corn oil by gavage. The Committee considered the result of the 13-week study to be valid because: (i) dietary administration is the more appropriate method of administration of compounds normally consumed in the diet; (ii) higher peak concentrations in tissues occur after administration of bolus doses by gavage than after dietary administration; (iii) micorencapsulation prevents loss due to volatilization of compounds such as furfural; (iv) microencapsulated furfural is rapidly released in an aqueous environment; and (v) corn oil is known to produce morphological changes in the livers of mice and rats when administered by gavage over a long period.
The results of an assay for unscheduled DNA synthesis in mice in vivo were reviewed by the Committee. This study, in which doses of up to 350 mg/kg bw were given, was particularly relevant since it addressed potential DNA repair in the cells in which tumours arose, namely hepatocytes. The negative results obtained in this assay were considered by the Committee to provide evidence that the liver tumours observed in the long-term study in mice were unlikely to have occurred through a genotoxic mechanism. The Committee considered that the concerns raised previously with respect to the liver tumours in mice were adequately addressed by this study and that a study of DNA binding was unnecessary.
The estimated daily intake of furfural from its use as a flavouring agent was determined from "poundage" data provided by the flavour industry. On the basis of these data, the estimated daily intake is 9 µg/kg bw in Europe and 8 µg/kg bw in the USA. The Committee was aware that intake of furfural from its use as a flavouring agent represents only a minor fraction of the total dietary intake of this substance.
The metabolism of 11 derivatives of furfuryl alcohol and furoic acid was considered by the Committee at its present meeting in the context of its evaluation of flavouring agents (see monograph on furfuryl alcohol and related flavouring agents). The derivatives were furfuryl acetate, furfuryl propionate, furfuryl pentanoate, furfuryl octanoate, furfuryl 3-methylbutanoate, methyl 2-furoate, propyl 2-furoate, amyl 2-furoate, hexyl 2-furoate, and octyl 2-furoate. Furfural, furfuryl alcohol, and these derivatives are metabolized to the same metabolite, furoic acid. Therefore, the Committee established a group ADI of 0–0.5 mg/kg bw for furfural and these structural analogues on the basis of the NOEL of 53 mg/kg bw per day in the 13-week study in rats and a safety factor of 100.
Buck, N. (2000) Release of furfural from microencapsulation through solvatin. Unpublished report from the Clinical Pharmacology Group, University of Southampton, United Kingdom. Submitted to WHO by the Flavor and Extract Manufacturers’ Association of the United States.
Edwards, A.J. (1999) An in vivo unscheduled DNA synthesis assay in the mouse with furfural. Unpublished report No. 3389/1/1/99 from BIBRA International, Carshalton, United Kingdom. Submitted to WHO by the Flavor and Extract Manufacturers’ Association of the United States.
Jonker, D. (2000a) Dose range finding study (14-day) with micro-encapsulated furfural in F344 rats. Unpublished report V98.1173 from TNO, Zeist, Netherlands. Submitted to WHO by the Flavor and Extract Manufacturers’ Association of the United States.
Jonker, D. (2000b) Amendment 1 to the TNO-report V99.520: Subchronic (13-week) oral toxicity study in rats with microencapsulated furfural. Unpublished report from TNO, Zeist, Netherlands, Submitted to WHO by the Flavor and Extract Manufacturers’ Association of the United States.
Jonker, D. (2000c) Sub-chronic (13-week) oral toxicity study in rats with micro-encapsulated furfural. Unpublished report V99.520 from TNO, Zeist, Netherlands. Submitted to WHO by the Flavor and Extract Manufacturers’ Association of the United States.
See Also: Toxicological Abbreviations Furfural (ICSC) Furfural (WHO Food Additives Series 30) Furfural (WHO Food Additives Series 42) FURFURAL (JECFA Evaluation) Furfural (IARC Summary & Evaluation, Volume 63, 1995)