Evaluation of Inner Exposure of Horses to Zearalenone (ZEN), Deoxynivalenol (DON) and Their Metabolites in Relation to Colic and Health-Related Clinical-Chemical Traits

Abstract

Mycotoxin contaminated feed has been associated with colic of horses caused by intestinal disorders. Whether such disease conditions alter the intestinal toxin metabolism and transfer across a compromised mucosal barrier is unknown. A screening approach was used to relate blood residue levels of DON, ZEN and their metabolites to the status of the horses (sick vs. healthy). A total of 55 clinically healthy horses from 6 different farms with varying feeding background served as control for sick horses (N = 102) hospitalized due to colic. ZEN, alpha-zearalenol (ZEL), beta-ZEL and DON were detectable in peripheral blood as indicators for the inner exposure with significant farm effects for alpha- and beta-ZEL. However, the levels in sick horses were similar to all farms. Moreover, the proportion of beta-ZEL of all detected ZEN metabolites as an indicator for the degree of metabolism of ZEN was not different for sick horses but differed amongst the control farms. Although the incidence of DON in blood was generally low and not significantly different amongst healthy and sick horses, the positive samples were nearly exclusively found in sick horses suggesting either a higher toxin transfer, an association of DON with the development of colic or a different feeding background.

Keywords: colic; deoxynivalenol; health; horse; intestinal disorders; metabolism; zearalenone.

Figure 1

Zearalenone (ZEN) (A), alpha-zearalenol (alpha-ZEL) (B) and beta-ZEL (C) concentrations in blood serum of horses, proportion of beta-ZEL of the sum of ZEN, alpha- and beta-ZEL (D) and serum deoxynivalenol (DON) concentrations (E) in horses hospitalized for colic (Farm 1, n = 102) and of clinically healthy horses from 6 different farms (Farm 2 to 7, n = 7–12). Boxes represent the span between the 25th and 75th percentile, the horizontal line within the boxes the median and the whiskers the range between minimum and maximum values. ab, distributions not sharing similar superscripts differ significantly (p < 0.05). Limits of detection (LOD) and of quantification (LOQ) are shown in (F) for ZEN, alpha-ZEL, beta-ZEL, zearalanone (ZAN), alpha-zearalanol (alpha-ZAL), beta-ZAL, DON and de-epoxy-DON (DOM-1).

Figure 2

Total protein and albumin concentrations (A,B), kynurenine (Kyn) to tryptophan (Trp) ratio (C), aspartate amino transferase (D), gamma-glutamyl transferase (E) and glutamate dehydrogenase (F) activities in blood of horses hospitalized for colic (Farm 1, n = 102) and of clinically healthy horses from 6 different farms (Farm 2 to 7, n = 7–12). Boxes represent the span between the 25th and 75th percentile, the horizontal line within the boxes the median and the whiskers the range between minimum and maximum values except for the Kyn to Trp ratio, where the boxes represent the ± range of the standard deviation and the horizontal line the mean value. ab distributions or mean values not sharing similar superscripts differ significantly (p < 0.05). Reference ranges (minimum-maximum) or maximum reference values [36,37] are indicated by horizontal red broken lines.

Figure 3

Serum albumin concentration in dependence on total protein content in serum (y = 20.5 + 0.27·x, r² = 0.27, p < 0.001, residual standard deviation = 3.0 g/L, N = 153) (A). Associations between aspartate amino transferase (AST) activity and kynurenine (Kyn) to tryptophan (Trp) ratio in serum in relation to the reference value for AST and to the highest Kyn to Trp ratio observed in healthy horses (broken lines) (B). Farm 1 , 2 , 3 , 4 , 5 , 6 , 7 .

Figure 4

Visualization of the results of the principal component analysis. Shown are the projection of the variables and of the status (healthy or intestine disorders) (A), and of the cases (individual horses) (B), respectively, into the space spanned between the first two components (PC or factors). Abbreviations for serum variables: GGT = gamma-glutamyl-transferase; GLDH = glutamate dehydrogenase; AST = aspartate aminotransferase; Prot = total protein; Alb = albumin; Trp = tryptophan; Kyn = Kynurenine; Kyn_Trp = ratio between Kyn and Trp; a-ZEL = alpha-zearalenol (ZEL); b-ZEL = beta-ZEL; ZEN = zearalenone; b_prop = proportion of b-ZEL of the sum of ZEN, a- and b-ZEL; DON = deoxynivalenol. Farm , 2 , 3 , 4 , 5 , 6 , 7 .

Figure 5

Time profiles of zearalenone (ZEN, in blue) and beta-zearalenol (beta-ZEL, in red) concentrations in blood of horses fed a ration containing 19 µg ZEN and 512 µg DON/kg feed corresponding to an exposure of 53 (±4) ng ZEN/kg body weight (bw) and 1420 (±3) ng DON/kg bw, respectively (A). Exposure was calculated based on this single morning meal given at −60 min as hay, and at 0 min as concentrate feed. Blood was sampled before offering these feed components and frequently until 300 min after offering the concentrate feed proportion (n = 5). Only values higher than the limits of quantification (LOQ) were plotted. Slopes of the linear regressions were not significantly different from zero (p > 0.05). Tryptophan (Trp) (B) and kynurenine (Kyn) (C) concentrations and the resulting molar ratio of Kyn to Trp (D) in blood of the same horses. Squares represent the lsmeans and whiskers the standard errors of the lsmeans (n = 5); a–e lsmeans not sharing similar superscripts differ significantly (p < 0.05).