A method for estimating the outer exposure of dairy cows to deoxynivalenol (DON) and zearalenone (ZEN) as a precondition for risk assessment based on inner exposure with toxin residue levels in blood and urine as indicators.

DON and ZEN residues in the blood and urine of dairy cows can be used to predict the outer exposure to DON and ZEN expressed per kilogram diet for a risk evaluation based on comparisons to critical dietary concentrations. This method was used to evaluate the exposure of dairy cows from 12 farms located in Brandenburg, Germany, fed rations with unknown DON and ZEN concentrations (N = 244). The corresponding diet concentrations predicted by different methods from analyzed blood and urine samples varied significantly amongst farms from 0 to 1.6 mg/kg for DON and 0 to 3.0 mg/kg for ZEN at a reference dry matter content of 88% but independently of lactational state (post-partum vs. early lactation). This significant variation was noticed below the critical dietary DON concentration of 5 mg/kg, while the ZEN concentration in one farm exceeded the critical ZEN level of 0.5 mg/kg markedly. Predicted DON concentrations of rations increased with the proportion of maize silage, while the high ZEN concentration found in one farm was most likely related to a higher proportion of sugar beet pulp supposedly highly contaminated by ZEN. Exceeding the critical dietary ZEN concentration and significant variations in DON contents below the critical level was not related to performance, reproductive performance, and health-related traits of cows. For a more consistent evaluation of possible associations between the inner exposure of cows to DON and ZEN, more frequent longitudinal observations of both mycotoxin residue levels and performance and health traits are required.

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

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.

Effects of the mycotoxin metabolite de-epoxy-deoxynivalenol (DOM-1) on embryo development and sperm motility in cattle.

Contamination of animal feed with Fusarium spp results in accumulation of mycotoxins including deoxynivalenol. In animals, deoxynivalenol is metabolized to de-epoxy deoxynivalenol (DOM-1), which is generally considered to be a non-toxic metabolite; however, recent studies demonstrated that DOM-1 can reduce steroid production and induce apoptosis in the bovine ovary. The objectives of this study were to assess the effects of DOM-1 on applied aspects of reproductive function in cattle, specifically sperm function and embryo development in vitro and follicle growth and superovulatory responses in vivo. The effect of naturally contaminated feed on superovulatory responses was assessed; a dose of 6 ppm deoxynivalenol increased blood DOM-1 concentrations to 20 ng/ml, but this did not alter the number of viable embryos recovered on day 7. However, intrafollicular injection of DOM-1 (100 ng/ml) directly into the growing dominant follicle resulted in cessation of follicular growth over the subsequent 3 days. Treatment with DOM-1 reduced motility of bull spermatozoa over a 10-h period in vitro. Addition of DOM-1 to oocytes in vitro during IVM did not alter rates of cumulus expansion and nuclear maturation, but treatment during IVF reduced the rate of blastocyst formation. These data illustrate that DOM-1 is more biologically active than previously thought and negatively impacted reproductive outcomes in cattle.

Maternal Exposure Results in Long-Term Deoxynivalenol Persistence in Piglets' Plasma and Modulates the Immune System.

Deoxynivalenol (DON)-contaminated feed represents a serious problem for pigs due to their high sensitivity to its toxicological effects. The aim of the present study was to evaluate the impact of intrauterine DON exposure on the immune system of piglets. Pure DON was intravenously administered to sows at the end of gestation (during the last 2-3 days of gestation, one dose of 300 µg per day). The plasma concentration of DON was analyzed using liquid chromatography combined with high-resolution Orbitrap-based mass spectrometry (LC-MS/MS (HR)) and selected immune parameters were monitored six times in piglets from birth to 18 weeks. DON was found in the plasma of 90% of newborn piglets at a mean concentration of 6.28 ng/mL and subsequently, at one, three, and seven weeks after birth with decreasing concentrations. Trace amounts were still present in the plasma 14 weeks after birth. Flow cytometry revealed a significant impact of DON on T lymphocyte subpopulations during the early postnatal period. Lower percentages of regulatory T cells, T helper lymphocytes, and their double positive CD4+CD8+ subset were followed by increased percentages of cytotoxic T lymphocytes and γδ T cells. The capacity to produce pro-inflammatory cytokines was also significantly lower after intrauterine DON exposure. In conclusion, this study revealed a long-term persistence of DON in the plasma of the piglets as a consequence of short-term intrauterine exposure, leading to altered immune parameters.

Toxicokinetic profile of fusarenon-X and its metabolite nivalenol in the goat (Capra hircus).

The main aim of this research was to evaluate the toxicokinetic characteristics of fusarenon-X (FX) and its metabolite, nivalenol (NIV), in goats. The amounts of FX and NIV in post-mitochondrial (S-9), microsomal and cytosolic fractions of diverse tissues of the goat were also investigated. FX was intravenously (iv) or orally (po) administered to goats at dosages of 0.25 and 1 mg/kg bw, respectively. The concentrations of FX and NIV in plasma, feces and urine were quantified by liquid chromatography tandem-mass spectrometry (LC-ESI-MS/MS). The concentrations of FX in plasma were quantified up to 8 h with both routes of administration. A large amount of NIV (metabolite) was quantifiable in plasma, urine and feces after both administrations. The Cmax value of FX was 413.39 ±â€¯206.84 ng/ml after po administration. The elimination half-life values were 1.64 ±â€¯0.32 h and 4.69 ±â€¯1.25 h after iv and po administration, respectively. In vitro experiments showed that the conversion FX-to-NIV mainly occurs in the liver microsomal fraction. This is the first study that evaluates the fate and metabolism of FX in ruminant species.

Prediction of deoxynivalenol toxicokinetics in humans by in vitro-to-in vivo extrapolation and allometric scaling of in vivo animal data.

Deoxynivalenol (DON) is the most prevalent mycotoxin in cereals worldwide. It can cause adverse health effects in humans and animals, and maximum levels in food and feed have been implemented by food authorities based on risk assessments derived from estimated intake levels. The lack of human toxicokinetic data such as absorption, distribution, and elimination characteristics hinders the direct calculation of DON plasma levels and exposure. In the present study, we have, therefore, used in vitro-to-in vivo extrapolation of depletion constants in hepatic microsomes from different species and allometric scaling of reported in vivo animal parameters to predict the plasma clearance [0.24 L/(h × kg)] and volume of distribution (1.24 L/kg) for DON in humans. In addition, we have performed a toxicokinetic study with oral and intravenous administration of DON in pigs to establish benchmark parameters for the in vitro extrapolation approach. The determined human toxicokinetic parameters were then used to calculate the bioavailability (50-90%), maximum concentration, and total exposure in plasma, and urinary concentrations under consideration of typical DON levels in grain-based food products. The results were compared to data from biomonitoring studies in human populations.

Detoxification of Fusarium-contaminated maize with sodium sulphite - in vivo efficacy with special emphasis on mycotoxin residues and piglet health.

A feeding experiment with piglets was performed to examine the efficacy of a wet preservation of Fusarium (FUS)-contaminated maize with sodium sulphite (SoS) based on deoxynivalenol (DON) and zearalenone (ZEN) residue levels in urine, bile and liquor and health traits of piglets. For this purpose, 80 castrated male piglets (7.57 ± 0.92 kg BW) were assigned to four treatment groups: CON- (control diet, with 0.09 mg DON and <0.01 mg ZEN/kg diet), CON+ (diet CON-, wet-preserved with 5 g SoS/kg maize; containing 0.05 mg DON and <0.01 mg ZEN/kg diet), FUS- (diet with mycotoxin-contaminated maize; containing 5.36 mg DON and 0.29 mg ZEN/kg diet), and FUS+ (diet FUS-, wet-preserved with 5 g SoS/kg maize; resulting in 0.83 mg DON and 0.27 mg ZEN/kg diet). After 42 d, 40 piglets (n = 10 per group) were sampled. A clear reduction of DON levels by approximately 75% was detected in all specimens of pigs fed diet FUS+. ZEN was detected in all urine, bile and liquor samples, while their metabolites were only detectable in urine and bile. Additionally, their concentrations were not influenced by SoS treatment. Among the health-related traits, feeding of FUS diets increased the total counts of leukocytes and segmented neutrophil granulocytes irrespective of SoS treatment. SoS treatment increased the total blood protein content slightly with a similar numerical trend in albumin concentration. These effects occurred at an obviously lower level in FUS-fed groups. Moreover, SoS treatment recovered the reduction of NO production induced by feeding diet FUS- indicating an effect on the redox level. As this effect only occurred in group FUS+, it is obviously related to the adverse effects of the Fusarium toxins. In conclusion, treatment of FUS-contaminated maize with SoS decreased the inner exposure with DON as indicated by the lower DON levels in various piglet specimens. However, health-related traits did not consistently reflect this decreased exposure.

Effects of oral exposure to sodium sulphite-treated deoxynivalenol (DON)-contaminated maize on performance and plasma concentrations of toxins and metabolites in piglets.

The objective of the present study was to demonstrate the efficiency of the decontamination process applied to deoxynivalenol (DON)-contaminated maize by sodium sulphite (Na2SO3) treatment in vivo. Additionally, in vitro characterisation of the toxicity of the DON sulphonates (DONS 1, 2 and 3 denote structurally different forms), the resulting DON metabolites, on peripheral blood mononuclear cells (PBMC) should substantiate the inactivation of DON. In a piglet experiment, both DON-contaminated maize and -uncontaminated control maize either untreated (DON-, CON-) or Na2SO3-treated (DON+, CON+) were mixed into feed and fed for 42 d starting from weaning. The results showed that feed intake and daily weight gain of animals fed DON- were significantly lower compared to animals fed CON- and CON+, whereas group DON+ reached the control level or even exceeded it. The feed-to-gain ratio was unaffected (p = 0.45). Furthermore, DON concentrations in plasma markedly reflected the diets' DON concentrations. These were < 0.1, < 0.1, 5.4 and 0.8 mg/kg feed for CON-, CON+, DON- and DON+, and amounted to 0.3, 0.4, 33.0 and 9.3 ng/ml in plasma, respectively. Whereas DONS 2 and 3 were detected in the DON+ diet, only DONS 2 was recovered in plasma. Regarding the toxicity of DONS, no or much lower toxicity was found compared to DON. DONS 1 and Na2SO3 did not affect the viability of PBMC. At 32.71µM DONS2 the viability was reduced by 50% and thus this compound was less toxic than DON by a factor of 73. Consequently, wet preservation of maize with Na2SO3 was an effective tool to avoid the adverse effects of DON on performance of piglets.

Plasma kinetics and matrix residues of deoxynivalenol (DON) and zearalenone (ZEN) are altered in endotoxaemic pigs independent of LPS entry site.

This study aimed to investigate a potential modulatory effect of E. coli lipopolysaccharide (LPS) on the kinetics of deoxynivalenol (DON) and zearalenone (ZEN) after pre- or post-hepatic LPS administration to unravel the putative role of the liver. Fifteen barrows were fed a diet containing mycotoxin-contaminated maize (4.59 mg DON/kg feed, 0.22 mg ZEN/kg feed) for 29 days and equipped with pre-hepatic catheters (portal vein, "po") and post-hepatic catheters (jugular vein, "ju"), facilitating simultaneous infusion of LPS ("LPS group", 7.5 µg/kg body weight) or 0.9% sterile NaCl solution (control, "CON group", equivolumar to LPS group) and blood sampling. This resulted in three infusion groups, depending on infusion site: CONju-CONpo, CONju-LPSpo, and LPSju-CONpo. On day 29, pigs were fed their morning ration (700 g/pig) (-15 min), and blood samples were collected at regular intervals relative to infusion start. At 195 min, pigs were sacrificed and bile, urine, liquor, and liver samples collected. DON concentrations in jugular and portal blood decreased in both LPS-infused groups, whereas the ZEN concentrations increased, regardless of the treatment site. In liver tissue, a decrease of both toxin concentrations was observed in endotoxaemic pigs as well as a drop in hepatic conjugation, regardless of LPS entry site. In contrast to our hypothesis, DON and ZEN were not differently altered depending on the LPS-entry site. Neither the absorption nor the accumulation of DON and ZEN in different tissues differed significantly between animals which were infused with LPS via either the jugular or portal vein.

Effects of deoxynivalenol (DON), zearalenone (ZEN), and related metabolites on equine peripheral blood mononuclear cells (PBMC) in vitro and background occurrence of these toxins in horses.

Both deoxynivalenol (DON), zearalenone (ZEN), and their metabolites are known to modulate immune cells in various species whereby viability and proliferation are influenced. Such effects were rarely examined in horses. Therefore, one aim of the present study was to titrate the inhibitory concentrations of DON, 3-acetyl-DON (3AcDON), de-epoxy-DON (DOM-1), ZEN, and α- and ß-zearalenol (ZEL) at which viability and proliferation of equine PBMC were reduced by 50 % (IC50) and 10 % (IC10) in vitro. For evaluation of practical relevance of the in vitro findings, a further aim was to screen horses for the background occurrence of DON, ZEN, and their metabolites in systemic circulation and to relate toxin residues both to the inhibitory toxin concentrations and to hematological and clinical-chemical characteristics.The IC50 (µM) for DON, 3AcDON, ß-ZEL, α-ZEL, and ZEN were determined at 3.09, 25.90, 75.44, 97.44, and 98.15 in unstimulated cells, respectively, while in proliferating cells, the corresponding IC50 values were 0.73, 6.89, 45.16, 75.96, and 82.51. Neither viability nor proliferation was influenced by DOM-1 up to a concentration of 100 µM.The in vivo screening (N = 49) revealed the occurrence of ZEN (N = 24), α-ZEL (N = 3), ß-ZEL (N = 37), DON, and DOM-1 (N = 2). The detected concentrations were much lower than the corresponding IC50 while the IC10 of DON and ß-ZEL for proliferating PBMC corresponded to approximately 26 and 35 ng/mL which might be relevant when contaminated diets are fed.Clinical-chemical and hematological traits were not related to mycotoxin residue levels excepting blood urea nitrogen which was positively correlated to the sum of ß-ZEL, α-ZEL, and ZEN concentration. Whether this reflects simply the feeding history of the horses or renal failures giving rise to a prolonged half-life of the toxins needs to be clarified further.

Prevalence of Selenium, T-2 Toxin, and Deoxynivalenol in Kashin-Beck Disease Areas in Qinghai Province, Northwest China.

The aim of this study was to determine the levels of selenium, T-2 toxin, and deoxynivalenol (DON) contamination in Kashin-Beck disease (KBD) areas and provide information for understanding the high prevalence of KBD in Qinghai Province. A total of 183 subjects were chosen in a KBD-prevalent county (Guide County) and a non-KBD county (Huangzhong County) in Qinghai Province, northwestern China, and the samples of wheat flour, soil, drinking water and blood, urine, and hair of children were collected from these residents. The selenium concentrations from all these sources were determined using atomic fluorescence spectrophotometry. The levels of T-2 toxin and DON contamination in the wheat flour samples were assayed using HPLC-MS/MS. The average selenium content in the soil, drinking water, and wheat flour samples from KBD areas were 26.93 ± 10.06 µg/kg, 0.097 ± 0.038 µg/L, and 9.50 ± 7.17 µg/kg, respectively. Among these, the selenium levels in the drinking water and wheat flour samples from the KBD endemic county were significantly lower than those from the non-KBD county. For the selenium nutrient status, only the hair selenium concentration of children from the KBD endemic county was significantly lower than that from the non-KBD county. The contents of T-2 toxin in all wheat samples were below the detection limit (0.4 µg/kg). The levels of DON contamination in wheat flour samples from KBD and non-KBD children's households within the KBD endemic county were relatively higher, with average levels of 302 ± 49 and 280 ± 48 µg/kg, respectively. The DON level of wheat flour samples from the children's households in the non-KBD county was significantly lower than that from the KBD endemic county. These results suggest that the lower selenium status in Guide County still remains. While the selenium nutritional status of the local children has improved to some extent, partly due to the introduction of food produce from nonlocal areas. DON contamination in the wheat flour may be involved in the fluctuating high prevalence rates of KBD in children in the KBD endemic Guide County in Qinghai Province.

Risk Assessment of Deoxynivalenol by Revisiting Its Bioavailability in Pig and Rat Models to Establish Which Is More Suitable.

Due to its toxic properties, high stability, and prevalence, the presence of deoxynivalenol (DON) in the food chain is a major threat to food safety and therefore a health risk for both humans and animals. In this study, experiments were carried out with sows and female rats to examine the kinetics of DON after intravenous and oral administration at 100 µg/kg of body weight. After intravenous administration of DON in pigs, a two-compartment model with rapid initial distribution (0.030 ± 0.019 h) followed by a slower terminal elimination phase (1.53 ± 0.54 h) was fitted to the concentration profile of DON in pig plasma. In rats, a short elimination half-life (0.46 h) and a clearance of 2.59 L/h/kg were estimated by sparse sampling non-compartmental analysis. Following oral exposure, DON was rapidly absorbed and reached maximal plasma concentrations (Cmax) of 42.07 ± 8.48 and 10.44 ± 5.87 µg/L plasma after (t(max)) 1.44 ± 0.52 and 0.17 h in pigs and rats, respectively. The mean bioavailability of DON was 70.5% ± 25.6% for pigs and 47.3% for rats. In the framework of DON risk assessment, these two animal models could be useful in an exposure scenario in two different ways because of their different bioavailability.

Studies on the bioavailability of deoxynivalenol (DON) and DON sulfonate (DONS) 1, 2, and 3 in pigs fed with sodium sulfite-treated DON-contaminated maize.

Deoxynivalenol (DON) exposure of pigs might cause serious problems when critical dietary toxin concentrations are exceeded. As DON contamination of agricultural crops cannot be completely prevented, detoxification measures are needed. Wet preservation with sodium sulfite resulted in a significant DON reduction of naturally-contaminated maize in previous experiments. The preserved material had a characteristic DON sulfonates (DONS) pattern. DONS is known to be less toxic than DON but its stability was shown to depend on pH, which gives rise to the question if a back-conversion to DON occurs in vivo. Therefore, the toxicokinetics and bioavailability of DON and DONS were studied in pigs. After the administration of a single oral or intravenous bolus of DON or DONS, serial blood samples were collected and subsequently analyzed. DONS was not detectable after oral administration of DONS mixtures. The results showed further that the bioavailability of DONS as DON in pigs fed maize preserved wet with sodium sulfite was significantly decreased compared to untreated control maize (DON), indicating that DONS obviously did not convert back to DON to a large extent in vivo. Moreover, the fact that DONS was not detectable in systemic blood requires further investigations regarding their ingestive and/or metabolic fate.

Metabolic and hematological consequences of dietary deoxynivalenol interacting with systemic Escherichia coli lipopolysaccharide.

Previous studies have shown that chronic oral deoxynivalenol (DON) exposure modulated Escherichia coli lipopolysaccharide (LPS)-induced systemic inflammation, whereby the liver was suspected to play an important role. Thus, a total of 41 barrows was fed one of two maize-based diets, either a DON-diet (4.59 mg DON/kg feed, n = 19) or a control diet (CON, n = 22). Pigs were equipped with indwelling catheters for pre- or post-hepatic (portal vs. jugular catheter) infusion of either control (0.9% NaCl) or LPS (7.5 µg/kg BW) for 1h and frequent blood sampling. This design yielded six groups: CON_CONjugular­CONportal, CON_CONjugular­LPSportal, CON_LPSjugular­CONportal, DON_CONjugular­CONportal, DON_CONjugular­LPSportal and DON_LPSjugular­CONportal. Blood samples were analyzed for blood gases, electrolytes, glucose, pH, lactate and red hemogram. The red hemogram and electrolytes were not affected by DON and LPS. DON-feeding solely decreased portal glucose uptake (p < 0.05). LPS-decreased partial oxygen pressure (pO2) overall (p < 0.05), but reduced pCO2 only in arterial blood, and DON had no effect on either. Irrespective of catheter localization, LPS decreased pH and base-excess (p < 0.01), but increased lactate and anion-gap (p < 0.01), indicating an emerging lactic acidosis. Lactic acidosis was more pronounced in the group DON_LPSjugular-CONportal than in CON-fed counterparts (p < 0.05). DON-feeding aggravated the porcine acid-base balance in response to a subsequent immunostimulus dependent on its exposure site (pre- or post-hepatic).

Deoxynivalenol Impairs Weight Gain and Affects Markers of Gut Health after Low-Dose, Short-Term Exposure of Growing Pigs.

Deoxynivalenol (DON) is one of the major mycotoxins produced by Fusarium fungi, and exposure to this mycotoxin requires an assessment of the potential adverse effects, even at low toxin levels. The aim of this study was to investigate the effects of a short-term, low-dose DON exposure on various gut health parameters in pigs. Piglets received a commercial feed or the same feed contaminated with DON (0.9 mg/kg feed) for 10 days, and two hours after a DON bolus (0.28 mg/kg BW), weight gain was determined and samples of different segments of the intestine were collected. Even the selected low dose of DON in the diet negatively affected weight gain and induced histomorphological alterations in the duodenum and jejunum. The mRNA expression of different tight junction (TJ) proteins, especially occludin, of inflammatory markers, like interleukin-1 beta and interleukin-10 and the oxidative stress marker heme-oxigenase1, were affected along the intestine by low levels of DON in the diet. Taken together, our results indicate that even after low-level exposure to DON, which has been generally considered as acceptable in animal feeds, clinically-relevant changes are measurable in markers of gut health and integrity.

The fate and tissue disposition of deoxynivalenol in broiler chickens.

To evaluate the fate of deoxynivalenol (DON) in broilers, DON was administered either intravenously or orally to broilers at a dose of 1 mg/kg BW. Concentrations of DON in plasma were measurable up to 4 hr and 2 hr after intravenous and oral administration, respectively. Following intravenous administration, the values for the elimination half-life, the volume of distribution and the clearance were 1.25 ± 0.25 hr, 7.55 ± 2.03 l/kg and 4.16 ± 0.42 l/hr/kg, respectively. The oral bioavailability was 15.46 ± 4.02%. DON was detectable in all tissues examined after oral administration. These results suggest that DON is able to penetrate into the various tissues in broilers, though poorly absorbed from their gastrointestinal tract.

Chronic exposure to deoxynivalenol has no influence on the oral bioavailability of fumonisin B1 in broiler chickens.

Both deoxynivalenol (DON) and fumonisin B1 (FB1) are common contaminants of feed. Fumonisins (FBs) in general have a very limited oral bioavailability in healthy animals. Previous studies have demonstrated that chronic exposure to DON impairs the intestinal barrier function and integrity, by affecting the intestinal surface area and function of the tight junctions. This might influence the oral bioavailability of FB1, and possibly lead to altered toxicity of this mycotoxin. A toxicokinetic study was performed with two groups of 6 broiler chickens, which were all administered an oral bolus of 2.5 mg FBs/kg BW after three-week exposure to either uncontaminated feed (group 1) or feed contaminated with 3.12 mg DON/kg feed (group 2). No significant differences in toxicokinetic parameters of FB1 could be demonstrated between the groups. Also, no increased or decreased body exposure to FB1 was observed, since the relative oral bioavailability of FB1 after chronic DON exposure was 92.2%.

Efficacy of active carbon towards the absorption of deoxynivalenol in pigs.

In order to assess the in vivo efficacy of mycotoxin binders, specific toxicokinetic parameters should be measured according to European guidelines. For this purpose, an absorption model in pigs is described with emphasis on absorption kinetics. Pigs received a single oral bolus of the mycotoxin deoxynivalenol alone or in combination with active carbon (applied as mycotoxin binder). After administration of deoxynivalenol alone, significant plasma amounts of deoxynivalenol were detected and kinetic parameters were calculated using a one compartmental model. Activated carbon completely prevented the absorption of deoxynivalenol as no plasma amounts could be detected.

Development and validation of an LC-MS/MS method for the toxicokinetic study of deoxynivalenol and its acetylated derivatives in chicken and pig plasma.

This study aims to develop an LC-MS/MS method allowing the determination of 3-acetyl-deoxynivalenol, 15-acetyl-deoxynivalenol, deoxynivalenol and its main in vivo metabolite, deepoxy-deoxynivalenol, in broiler chickens and pigs. These species have a high exposure to these toxins, given their mainly cereal based diet. Several sample cleanup strategies were tested and further optimized by means of fractional factorial designs. A simple and straightforward sample preparation method was developed consisting out of a deproteinisation step with acetonitrile, followed by evaporation of the supernatant and reconstitution in water. The method was single laboratory validated according to European guidelines and found to be applicable for the intended purpose, with a linear response up to 200ngml(-1) and limits of quantification of 0.1-2ngml(-1). As a proof of concept, biological samples from a broiler chicken that received either deoxynivalenol, 3- or 15-acetyl-deoxynivalenol were analyzed. Preliminary results indicate nearly complete hydrolysis of 3-acetyl-deoxynivalenol to deoxynivalenol; and to a lesser extent of 15-acetyl-deoxynivalenol to deoxynivalenol. No deepoxy-deoxynivalenol was detected in any of the plasma samples. The method will be applied to study full toxicokinetic properties of deoxynivalenol, 3-acetyl-deoxynivalenol and 15-acetyl-deoxynivalenol in broiler chickens and pigs.

Residues of deoxynivalenol (DON) and its metabolite de-epoxy-DON in eggs, plasma and bile of laying hens of different genetic backgrounds.

In the present study, the potential for carry-over of deoxynivalenol (DON) into eggs and DON residues in plasma and bile of laying hens of different genetic backgrounds after long-term feeding trial was investigated. A total of 80, 23-week-old laying hens were assigned to a feeding trial with two diets, a control diet and a Fusarium toxin-contaminated diet (FUS) (0.4 and 9.9 mg DON kg(-1), respectively). In the 60th week of hen's life, 10 eggs from each group were collected. In the 70th week of hen's life, all hens were slaughtered and samples of blood and bile were collected. The samples were analysed by liquid chromatography tandem mass spectrometry (LC-MS/MS) for DON and de-epoxy-DON. DON was only detected in samples of hens which fed the FUS diet while none of the samples analysed had detectable levels of de-epoxy-DON. In plasma and bile samples, DON levels ranged from 0.2 to 0.6 ng ml(-1) and from 1.8 to 4.1 ng ml(-1), respectively. DON levels in egg yolk and albumen ranged between 0.0-0.46 ng g(-1) and 0.0-0.35 ng g(-1), respectively, corresponding to carry-over rates of DON into eggs from 0.0 to 0.000016. Moreover, no differences in DON levels or carry-over rates were noticed between the two tested breeds. These results show that very low levels of DON were transferred into eggs and indicate that although eggs could contribute to human exposure to DON, the levels are very low and insignificant.