Validation of fumonisin biomarkers in F344 rats

Abstract

Fumonisins (FNs) are ubiquitous contaminants of cereal grains. Fumonisin B(1) (FB(1)) was linked to several animal and human diseases. To validate FB(1) biomarkers for studying human disease risks, F344 rats were administered by gavage with either a single dose of 0, 10 or 25 mg FB(1)/kg body weight (BW) or repeated doses of 0, 1.0, or 2.5 mg FB(1)/kg BW/day for 5 weeks. FB(1) excretion and FB(1)-induced metabolic alterations of sphingolipids in rat urine, feces and serum were assessed. Dose-dependent urinary and fecal excretion of free FB(1) were found in both single-dose- and repeat-dose-treated rats. In the single-dose study, urinary sphinganine (Sa) to sphingosine (So) ratio (Sa/So) reached a maximum at day 7 for the high-dose group and at day 5 for the low-dose group, whereas serum Sa/So showed only marginal changes. In the repeat-dose study, urinary Sa/So was persistently elevated at 2 weeks, while serum Sa/So was unchanged. Time course changes of sphinganine 1-phosphate (SaP) and sphingosine 1-phosphate (SoP) were also examined. Although serum Sa/So and SaP/SoP ratios showed no signs of time- or dose-dependent changes, a 10-fold increase in urinary SaP/SoP was observed, suggesting that urinary SaP/SoP is a more sensitive biomarker for FB(1) exposure. The accumulation of SaP and SoP was evident in the time course of SaP/Sa and SoP/So, which may reflect activity changes of enzymes closely related to the metabolism and catabolism of SaP and SoP. These results provide concrete evidence towards the practical use of excreted FB(1), Sa/So and SaP/SoP as biomarkers of exposure to FNs.

Fig. 1

Chemical structure of FB₁ and the major sphingolipid metabolites affected.

Fig. 2

Experimental design for the validation of sphingolipid metabolites as FB₁ biomarkers in F344 rats.

Fig. 3

Representative LC/MS total ion chromatograms of a standard (A) containing C17SaP, SoP, SaP, So, Sa, and C20So (5 pmol injection each analyte), and urine extracts from a control rat (B), rats receiving 1.0 (C) and 2.5 (D) mg/kg BW/day of FB₁.

Fig. 4

Urinary levels of free FB₁ in rats receiving a single-dose (A) or repeat-dose (B) of FB₁, and fecal levels of free FB₁ in rats receiving a single-dose (C) or repeat-dose (D) of FB₁. Asterisks indicate time points where the results were significantly different from the low-dose group in the single-dose study or the control group in the repeat-dose study (* and ** for P<0.05 and P<0.01, respectively).

Fig. 5

Urinary ratios of Sa/So (A) and SaP/SoP (B) in rats receiving single-dose of 10 or 25 mg/kg BW FB₁. The asterisks (* and **) indicate time points where the results of 25 and 10 mg/kg BW doses were significantly different (P<0.05 and P<0.01, respectively).

Fig. 6

Serum ratios of Sa/So (A) and SaP/SoP (B) in rats receiving single-dose of 10 or 25 mg/kg BW FB₁.

Fig. 7

Urinary ratios of Sa/So (A), SaP/SoP (B), SaP/Sa (C), and SoP/So (D) in rats receiving repeat-dose of 0.0, 1.0, or 2.5 mg/kg BW/day FB₁. The asterisks (* and **) indicate time points where the results were significantly different from the control group with P<0.05 and P<0.01, respectively.

Fig. 8

Serum ratios of Sa/So (A), SaP/SoP (B), SaP/Sa (C), and SoP/So (D) in rats receiving repeat-dose of 0.0, 1.0 or 2.5 mg/kg BW/day FB₁. The asterisks (* and **) indicate time points where the results were significantly different from the control group with P<0.05 and P<0.01, respectively.

Fig. 9

Disruption of de novo sphingolipid biosynthetic and turnover pathways by FB₁ treatment.