Efficacy of Fumonisin Esterase in Piglets as Animal Model for Fumonisin Detoxification in Humans: Pilot Study Comparing Intraoral to Intragastric Administration.

Fumonisins, a group of highly prevalent and toxic mycotoxins, are suspected to be causal agents of several diseases in animals and humans. In the animal feed industry, fumonisin esterase is used as feed additive to prevent mycotoxicosis caused by fumonisins. In humans, a popular dosage form for dietary supplements, with high patient acceptance for oral intake, is capsule ingestion. Thus, fumonisin esterase provided in a capsule could be an effective strategy against fumonisin intoxication in humans. To determine the efficacy of fumonisin esterase through capsule ingestion, two modes of application were compared using piglets in a small-scale preliminary study. The enzyme was administered intraorally (in-feed analogue) or intragastrically (capsule analogue), in combination with fumonisin B1 (FB1). Biomarkers for FB1 exposure; namely FB1, hydrolysed FB1 (HFB1) and partially hydrolysed forms (pHFB1a and pHFB1b), were measured both in serum and faeces using a validated liquid chromatography-tandem mass spectrometry (LC-MS/MS) method, and toxicokinetic parameters were calculated. Additionally, the serum sphinganine/sphingosine (Sa/So) ratio, a biomarker of effect, was determined using LC-MS/MS. A significantly higher Sa/So ratio was shown in the placebo group compared to both esterase treatments, demonstrating the efficacy of the esterase. Moreover, a significant decrease in serum FB1 area under the concentration-time curve (AUC) and an increase of faecal HFB1 AUC were observed after intraoral esterase administration. However, these effects were not observed with statistical significance after intragastric esterase administration with the current sample size.

The protective role of liver X receptor (LXR) during fumonisin B1-induced hepatotoxicity.

Fumonisin B1 (FB1), a congener of fumonisins produced by Fusarium species, is the most abundant and most toxicologically active fumonisin. FB1 causes severe mycotoxicosis in animals, including nephrotoxicity, hepatotoxicity, and disruption of the intestinal barrier. However, mechanisms associated with FB1 toxicity are still unclear. Preliminary studies have highlighted the role of liver X receptors (LXRs) during FB1 exposure. LXRs belong to the nuclear receptor family and control the expression of genes involved in cholesterol and lipid homeostasis. In this context, the toxicity of FB1 was compared in female wild-type (LXR+/+) and LXRα,ß double knockout (LXR-/-) mice in the absence or presence of FB1 (10 mg/kg body weight/day) for 28 days. Exposure to FB1 supplemented in the mice's drinking water resulted in more pronounced hepatotoxicity in LXR-/- mice compared to LXR+/+ mice, as indicated by hepatic transaminase levels (ALT, AST) and hepatic inflammatory and fibrotic lesions. Next, the effect of FB1 exposure on the liver transcriptome was investigated. FB1 exposure led to a specific transcriptional response in LXR-/- mice that included altered cholesterol and bile acid homeostasis. ELISA showed that these effects were associated with an elevated FB1 concentration in the plasma of LXR-/- mice, suggesting that LXRs participate in intestinal absorption and/or clearance of the toxin. In summary, this study demonstrates an important role of LXRs in protecting the liver against FB1-induced toxicity, suggesting an alternative mechanism not related to the inhibition of sphingolipid synthesis for mycotoxin toxicity.

Development and Validation of a UPLC-MS/MS and UPLC-HR-MS Method for the Determination of Fumonisin B1 and Its Hydrolysed Metabolites and Fumonisin B2 in Broiler Chicken Plasma.

A sensitive and specific method for the quantitative determination of Fumonisin B1 (FB1), its partially hydrolysed metabolites pHFB1a+b and hydrolysed metabolite HFB1, and Fumonisin B2 (FB2) in broiler chicken plasma using ultra-performance liquid chromatography combined with tandem mass spectrometry (UPLC-MS/MS) was developed. The sample preparation was rapid, straightforward and consisted of a deproteinization and phospholipid removal step using an Oasis® OstroTM 96-well plate. Chromatography was performed on an Acquity HSS-T3 column, using 0.3% formic acid and 10 mM ammonium formate in water, and acetonitrile as mobile phases. The MS/MS instrument was operated in the positive electrospray ionization mode and the two multiple reaction monitoring transitions were monitored for each component for quantification and identification, respectively. The method was validated in-house: matrix-matched calibration graphs were prepared and good linearity (r ≥ 0.99) was achieved over the concentration ranges tested (1-500 ng/mL for FB1 and FB2; 0.86-860 ng/mL for pHFB1a; 0.72-1430 ng/mL for pHFB1b and 2.5-2500 ng/mL for HFB1). Limits of quantification (LOQ) and detection (LOD) in plasma ranged between 0.72 to 2.5 ng/mL and 0.03 to 0.17 ng/mL, respectively. The results for the within-day and between-day precision and accuracy fell within the specified ranges. Moreover, the method was transferred to an UPLC high-resolution mass spectrometry (HR-MS) instrument in order to determine potential metabolites of HFB1, such as N-acyl-HFB1s and phase II metabolites. The method has been successfully applied to investigate the toxicokinetics and biotransformation of HFB1 in broiler chickens.

Determination of fumonisin B1 levels in body fluids and hair from piglets fed fumonisin B1-contaminated diets.

The levels of fumonisin B1 (FB1) residues in plasma, urine, feces and hair from 24 piglets fed FB1-contaminated diets containing 3.1, 6.1 or 9.0 µg FB1.g-1 for 28 days were determined using liquid chromatography coupled to mass spectrometry (LC-MS/MS). The levels of FB1 in plasma, urine, feces and pooled hair (n = 3) samples varied from 0.15 to 1.08 µg.L-1, 16.09-75.01 µg.L-1, 1.87-13.89 µg.g-1 and 2.08-8.09 ng.g-1, respectively. Significant correlations (r = 0.808-0.885; P < 0.001; N = 18) were found between FB1 intake and plasma FB1 on days 7, 14, 21 and 28. However, urinary FB1 correlated with FB1 intake only on days 7 and 14 (r = 0.561-572; P = 0.02; N = 18). A significant correlation (r = 0.509; P = 0.02; N = 24) was also found for the first time between FB1 in hair samples and FB1 intake. Plasma and urinary FB1 are good biomarkers of early exposure of pigs to low dietary FB1 levels, although plasma is recommended to assess prolonged exposure (>14 days). The possibility to evaluate hair as a biomarker of fumonisin exposure was established, although further studies are needed to provide physiologically based toxicokinetics of residual FB1 in the pig hair.

Relação entre níveis de fumonisina B1 e ácido fólico em farinha de milho e a concentração de ácido fólico no soro humano / Relationship between fumonisin B1 and folic acid levels in foods and the folic acid concentration in human serum

Neste trabalho foram determinados os níveis de ácido fólico e de fumonisina B1 (FB1) em farinha de milho consumida por 24 voluntários residentes em um campus universitário no estado de São Paulo, bem como sua relação com as concentrações de ácido fólico sérico nos indivíduos. As análises de ácido fólico e de FB1 em farinha de milho foram realizadas por cromatografia líquida de alta eficiência (CLAE), enquanto a determinação de ácido fólico sérico foi feita por kit de imunoensaio. Detectou-se a FB1 em 100% das amostras de farinha de milho, em níveis que variaram de 142 a 3.037 µg kg-1 (média: 738 ± 591 µg kg-1). As concentrações de ácido fólico nas amostras de farinha de milho ficaram entre < 0,3 µg kg-1 (limite de quantificação) e 1.705 µg kg-1, com média de 713 ± 435 µg kg-1, o que representa 47% do limite mínimo exigido pela Agência Nacional de Vigilância Sanitária (ANVISA) para farinhas de milho comercialmente disponíveis. Nas amostras de soro humano, os níveis de ácido fólico variaram de 6,7 a 24,0 ng mL-1 (média: 13,4 ± 5,4 ng mL-1). Não houve correlação (p < 0,05) entre os níveis de ácido fólico no soro dos indivíduos e as concentrações de FB1 ou ácido fólico nas amostras de farinha de milho. Outros estudos são necessários para estimar a ingestão total de FB1 por meio da dieta para averiguar os efeitos das fumonisinas sobre a absorção de ácido fólico nos indivíduos avaliados.(AU)

In the present study, folic acid and fumonisin B1 (FB1) levels were determined in corn flour consumed by 24 volunteers, residents in a university campus in São Paulo State, as well as its relationship with folic acid in serum of individuals. Analyses of folic acid and FB1 in corn flour were performed by high-performance liquid chromatography (HPLC), while the determination of folic acid in serum was accomplished using an immunoassay kit. FB1 was detected in 100% of corn samples, at levels ranging from 142 to 3,037 µg kg-1 (which means: 738 ± 591 µg kg-1). The concentrations of folic acid in corn flour samples ranged from < 0.3 µg kg-1 (limit of quantification) to 1,705 µg kg-1, with a mean of 713 ± 435 µg kg-1, which represents 47% of the minimum required by National Agency of Health Surveillance (ANVISA) for corn flour commercially available. The levels of folic acid in human serum samples ranged from 6.7 to 24.0 ng mL-1 (meaning: 13.4 ± 5.4 ng mL-1). No correlations were observed (p < 0.05) between the folic acid levels in serum of individuals and the concentrations of FB1 or folic acid in corn flour samples. Further studies are needed to estimate the total intake of FB1 in the diet to assess the effects fumonisins on the absorption of folic acid in the individuals evaluated.(AU)

Mitigation of Fumonisin Biomarkers by Green Tea Polyphenols in a High-Risk Population of Hepatocellular Carcinoma.

Green tea polyphenols (GTP) are highly effective in inhibiting a variety of tumorigenic effects induced by carcinogens. In this study we assessed GTP mitigation on biomarkers of fumonisin B1 (FB1), a class 2B carcinogen, in blood and urine samples collected from an intervention trial. A total of 124 exposed people were recruited and randomly assigned to low-dose (GTP 500 mg, n = 42), high-dose (GTP 1,000 mg, n = 41) or placebo (n = 41) for 3 months. After one-month of intervention, urinary FB1 was significantly decreased in high-dose group compared to that of placebo group (p = 0.045), with reduction rates of 18.95% in the low-dose group and 33.62% in the high-dose group. After three-month intervention, urinary FB1 showed significant decrease in both low-dose (p = 0.016) and the high-dose (p = 0.0005) groups compared to that of both placebo group and baseline levels, with reduction rates of 40.18% in the low-dose group and 52.6% in the high-dose group. GTP treatment also significantly reduced urinary excretion of sphinganine (Sa), sphingosine (So), and Sa/So ratio, but had no effect on serum Sa, So, and Sa/So ratio. Analysis with mixed-effect model revealed significant interactions between time and treatment effects of GTP on both urinary free FB1 levels and Sa/So ratios.

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%.

Mycotoxin exposure and infant and young child growth in Africa: what do we know?

INTRODUCTION: Infant and young child (IYC) growth impairment remains a public health problem in Africa partly because infants are exposed to staple foods (contaminated with mycotoxins) at an early age. Understanding the role of mycotoxins in IYC growth is vital, and this paper systematically reviews the available knowledge. METHODS: Studies were searched and included if they provided information on African IYC mycotoxin exposure rates and/or growth. Studies were excluded if subjects were older than 15 years, if they were animal studies or focusing on other mycotoxins. Relevant search words were included in search strings. Eight reviews were identified and reference lists scrutinised for additional studies. RESULTS: Ten studies were included; 8 focused on aflatoxin (AF), 2 on fumonisin (FB) and none on deoxynivalenol (DON) and zearalenone (ZEA). AF exposure prevalence reached 100% with levels at 40.4 pg/mg. AF was present in umbilical cords indicating that AF crosses the placenta. Maternal exposure levels were correlated with breast milk levels. The highest levels of serum AF (mean 32.8 pg/mg) were measured in Benin and Togo with 5.4% reaching levels higher than 200 pg/mg. At the end of weaning, children had similar prevalence and exposure levels as adults. RESULTS also indicated that infants with higher levels of maternal exposure had significantly lower height-for-age z-scores (HAZ scores), although there was no significant association between cord AF and infant HAZ scores or AF in cord blood and HAZ scores. Significantly higher mean maternal AF levels related to lower weight-for-age z-scores (WAZ scores) were reported, and infants with higher levels of maternal exposure had significantly lower WAZ scores that decreased over age. Cord AF levels had no effect on infant WAZ scores. One study investigated the association between FB and IYC growth and found that those with FB intakes greater than the provisional maximum tolerable daily intake were significantly shorter (1.3 cm) and lighter (328 g). No studies investigated the role of DON and ZEA. CONCLUSION: A limited number of epidemiological studies have been conducted, and available research indicates extreme exposures to AF. There are strong associations between AF exposure and stunting and wasting. However, more epidemiological research is urgently needed to understand the role of FB, DON and ZEA in IYC growth.

Calcium montmorillonite clay reduces AFB1 and FB1 biomarkers in rats exposed to single and co-exposures of aflatoxin and fumonisin.

Aflatoxins (AFs) and fumonisins (FBs) can co-contaminate foodstuffs and have been associated with hepatocellular and esophageal carcinomas in humans at high risk for exposure. One strategy to reduce exposure (and toxicity) from contaminated foodstuffs is the dietary inclusion of a montmorillonite clay (UPSN) that binds AFs and FBs in the gastrointestinal tract. In this study, the binding capacity of UPSN was evaluated for AFB1, FB1 and a combination thereof in Fischer 344 rats. Rats were pre-treated with different dietary levels of UPSN (0.25% or 2%) for 1 week. Rats were gavaged with a single dose of either 0.125 mg AFB1 or 25 mg FB1 per kg body weight and a combination thereof in the presence and absence of an aqueous solution of UPSN. The kinetics of mycotoxin excretion were monitored by analyzing serum AFB1 -albumin, urinary AF (AFM1) and FB1 biomarkers over a period of 72 h. UPSN decreased AFM1 excretion by 88-97%, indicating highly effective binding. FB1 excretion was reduced, to a lesser extent, ranging from 45% to 85%. When in combination, both AFB1 and FB1 binding occurred, but capacity was decreased by almost half. In the absence of UPSN, the combined AFB1 and FB1 treatment decreased the urinary biomarkers by 67% and 45% respectively, but increased levels of AFB1 -albumin, presumably by modulating its cytochrome metabolism. UPSN significantly reduced bioavailability of both AFB1 and FB1 when in combination; suggesting that it can be utilized to reduce levels below their respective thresholds for affecting adverse biological effects.

Biomonitoring of Fusarium spp. mycotoxins: perspectives for an individual exposure assessment tool.

Fusarium species are probably the most prevalent toxin-producing fungi of the northern temperate regions and are commonly found on cereals grown in the temperate regions of America, Europe and Asia. Among the toxins formed by Fusarium we find trichothecenes of the A-type or B-type, zearalenone, fumonisins or nivalenol. The current exposure assessment consists of the qualitative and/or quantitative evaluation based on the knowledge of the mycotoxin occurrence in the food and the dietary habits of the population. This process permits quantifying the mycotoxin dietary intake through deterministic or probabilistic methods. Although these methods are suitable to assess the exposure of populations to contaminants and to identify risk groups, they are not recommended to evaluate the individual exposition, due to a low accuracy and sensitivity. On the contrary, the use of biochemical indicators has been proposed as a suitable method to assess individual exposure to contaminants. In this work, several techniques to biomonitor the exposure to fumonisins, deoxynivalenol, zearalenone or T-2 toxin have been reviewed.