Aldo Rizzo

Presence and concentrations of the Fusarium-related mycotoxins beauvericin, enniatins and moniliformin in finnish grain samples.

Fusarium mycotoxins beauvericin, enniatins (A, A1, B, B1) and moniliformin were analysed in 38 Finnish grain samples (14 wheat, 22 barley, one rye, one oats) harvested in 2001–02. The contaminating Fusarium species were identified with the primer-specific polymerase chain reaction as well as with morphological studies. All the studied mycotoxins were found in the samples. Enniatins B and B1 were detected in all samples, and enniatin A, enniatin A1, beauvericin and moniliformin in 74, 95, 95 and 74% of the samples, respectively. There were higher concentrations of the mycotoxins analysed in 2001 compared with 2002. The highest levels of mycotoxins were detected in samples harvested late in the autumn after a long rainy period. Fusarium avenaceum was the most abundant Fusarium species in Finland during both years (0–29.5%) measured as infected kernels. A significant correlation was found between F. avenaceum contamination level and the concentration levels of enniatins B and B1, as well as moniliformin.

Determination of selected mycotoxins in mould cheeses with liquid chromatography coupled to tandem with mass spectrometry.

A simple and feasible method is described for analysing nine mycotoxins in cheese matrix. The method involves liquid extraction followed by high performance liquid chromatographic separation and mass spectrometric detection of the analytes, and allows the determination of aflatoxins B1, B2, G1, G2 and M1, ochratoxin A, mycophenolic acid, penicillic acid and roquefortine C simultaneously. Average recoveries of the mycotoxins from spiked samples at concentration levels of 5–200 µg kg−1 ranged from 96–143%. Within-day relative standard deviations at these concentration levels varied from 2.3–12.1%. The limit of quantification for aflatoxin M1 was 0.6 µg kg−1 and for the other compounds 5 µg kg−1. The method developed was applied for analysing these mycotoxins in blue and white mould cheeses purchased from Finnish supermarkets. Roquefortine C was detected in all of the blue mould cheese samples in concentrations of 0.8–12 mg kg−1. One blue cheese contained also 0.3 mg kg−1 mycophenolic acid. The other investigated mycotoxins were absent in the samples.

Trichothecenes, ochratoxin A and zearalenone contamination and Fusarium infection in Finnish cereal samples in 1998

The occurrences and concentrations of trichothecenes, ochratoxin A and zearalenone in Finnish cereal samples are presented in this study. Furthermore, infections by moulds, especially Fusarium contamination of grains in the same samples, are reported. In total 68 cereal samples, including 43 rye, 4 wheat, 15 barley and 6 oats samples, were collected after a cool and very rainy growing season in 1998. A gas chromatograph combined with a mass spectrometric detector was used for determination of seven different trichothecenes. A high performance liquid chromatograph with a fluorescence detector was used for ochratoxin A and zearalenone determination. For the identification of moulds, the grain samples were incubated and the moulds were isolated and identified by microscopy. The analytical methods were validated for mycotoxin analysis and they were found to be adequately reliable and sensitive. Heavy rainfalls in the summer and autumn of 1998 caused abundant Fusarium mould infection in Finnish cereals, particularly in rye. Fusarium avenaceum was the most common Fusarium species found in cereals. However, the mycotoxin concentrations were very low and only deoxynivalenol, nivalenol and HT-2 toxin were detected. Deoxynivalenol was detected in 54 samples in the concentration range 5-111μg/kg. Nivalenol and HT-2 toxin were detected in three and two samples, respectively, in the concentration range 10-20 μg/kg.

Real-time PCR for Quantification of Toxigenic Fusarium Species in Barley and Malt

A real-time PCR technique was applied for the quantification of trichothecene-producing Fusarium species (TMTRI assay) as well as the highly toxigenic Fusarium graminearum (TMFg12 assay) present in barley grain and malt. PCR results were compared to the amounts of trichothecenes detected in the samples to find out if the PCR assays can be used for trichothecene screening instead of expensive and laborious chemical analyses. DNA was extracted from ground kernels using a commercial DNA extraction kit and analysed in a LightCycler® system using specific primers and fluorogenic TaqMan probes. Both naturally and artificially contaminated grains were analysed. The TMTRI assay and the TMFg12 assay enabled the quantification of trichothecene-producing Fusarium DNA and F. graminearum DNA present in barley grain and malt samples, respectively. Both TaqMan assays were considered to be sensitive and reproducible. Linearity of the assays was 4–5 log units when pure Fusarium DNAs were tested. The amount of Fusarium DNA analysed with the TMTRI-trichothecene assay could be used for estimation of the deoxynivalenol (DON) content in barley grain. Furthermore, the TMFg12 assay for F. graminearum gave a good estimation of the DON content in north American barley and malt samples, whilst the correlation was poor among Finnish samples. DON content and the level of F. graminearum DNA were found to be naturally low in Finnish barleys.

RETRACTED: Antioxidant nutrients and mycotoxins

This article has been retracted at the request of the editor. Reason: This article contained extensive text that originally appeared in the Journal of Food Protection (FABIO GALVANO, ANDREA PIVA, ALBERTO RITIENI, and GIACOMO GALVANO published by the International Association for Food Protection. Dietary Strategies to Counteract the Effects of Mycotoxins: A Review. J. Food Prot. (2001) Vol. 64 (1): 120–131). Toxicology publishes only original work, and on the occasion when brief excerpts or quotes from other sources are used, this must be clearly indicated with full acknowledgement and written permission from the copyright holder.

Ochratoxin A in cereals, foodstuffs and human plasma

Aspergillus and Penicillium fungi contaminate cereals and foodstuffs, and can thus introduce ochratoxin A (OTA) into the food chain. In this work, five new isolates of Aspergillus: A. albertensis, A. auricomus, A. wentii, A. fumigatus and A. versicolor, were found to produce ochratoxin A. Data on the occurrence and the concentration levels of ochratoxin A in European food of vegetable and animal origin are reported. Furthermore, data on the concentration of ochratoxin A in blood of citizens of Western Europe are compared with those of some areas where Balkan endemic nephropathy is endemic. The results of the studies of Stoev and co-workers are reviewed and the possibility that OTA alone cannot be the cause of the Balkan endemic nephropathy is discussed.

Determination of Fusarium‐Mycotoxins Beauvericin and Enniatins with Liquid Chromatography–Tandem Mass Spectrometry (LC–MS/MS)

Abstract A liquid chromatography–mass spectrometric method was developed and validated to determine Fusarium mycotoxins beauvericin and enniatins (A, A1, B, B1) in grain samples. For the first time, a triple quadrupole mass spectrometer with multiple reaction monitoring was used for the analysis of these mycotoxins, which allowed the simultaneous structural identification of the analytes. The sample preparation involved direct purification of sample extracts with reversed‐phase–solid phase extraction, enabling the avoidance of time consuming and loss‐causing liquid–liquid extractions. The method validation included the determination of selectivity, repeatability, limit of detection, limit of quantification, recovery, and linearity. The developed method proved to be sensitive and repeatable for the analysis of the mycotoxins in grain matrix. Since the method was originally developed solely for beauvericin, the recoveries of some enniatins remained rather low, except at the lowest spiking level. This was due to the natural contamination of grain matrix with enniatins B and B1, which resulted in higher recoveries for these mycotoxins. The mean recoveries for beauvericin and enniatins A, A1, B, and B1 were 76–82%, 55–66%, 71–80%, 57–103%, and 68–116%, respectively. The calculated limits of quantification for beauvericin and enniatins A, A1, B, and B1 were 0.2, 0.2, 0.7, 0.9, and 1.5 µg/kg, respectively.

Genetic variation, real-time PCR, metabolites and mycotoxins ofFusarium avenaceum and related species.

In this paper the latest studies dealing with genetic variation and mycotoxins ofF. avenaceum and related species are reviewed and compared to the data from chromatographic image analyses. Forty-three European strains ofFusarium avenaceum and related species were classified by chromatographic image analysis on full chromatographic matrices. The results were in most cases in agreement with those from morphological and molecular analyses and supported the separation betweenF. avenaceum, F. arthrosporioides andF. tricinctum and betweenF. avenaceum groups I and II. The mycotoxin profiles of the FinnishF. avenaceum, F. arthrosporioides andF tricinctum strains were very similar to each other. Moniliformin and enniatins were the main mycotoxins produced. A fluorogenic TaqMan PCR assay (qPCR) was used for the detection ofF. avenaceum/ F. arthrosporioides DNA in Finnish barley and wheat. The qPCR results obtained from grain samples were compared to mycotoxin levels. A correlation was found betweenF. avenaceum/F. arthrosporioides DNA and moniliformin (MON) and enniatin (ENNs) levels in barley. A correlation was also found between the combinedF. avenaceum/F. arthrosporioides/F. tricinctum contamination and MON and ENNs levels in barley in 2002, but not in 2003. This was probably due to the higher MON and ENNs levels in 2002 than in 2003. It was possible to use the DNA levels ofF. avenaceum/F. arthrosporioides to distinguish between most barley samples containing high amounts of MON and ENNs from those containing low levels of the mycotoxins.

The characterization of trichothecenes as their heptafluorobutyrate esters by negative-ion chemical ionization tandem mass spectrometry

Abstract Negative-ion chemical ionization (NCI) and collisionally activated mass spectra are presented for heptafluorobutyryl (HFB) esters of eight trichothecenes. The ion-source conditions have a dramatic effect on the NCI spectra and only at low source temperatures can intense parent ions, i.e., M − for bis- and tris-HFB esters (neosolaniol, HT-2 Toxin, monoacetosyscirpenol, fusarenon-X and deoxynivalenol) and [M-HF] − for mono-HFB esters (T-2 Toxin, Iso-T-2 Toxin, and diacetoxyscirpenol), be generated. High selectivity and a sensitivity down to picogram (0.2-2pg) levels were achieved with gas chromatography/NCI tandem mass spectrometry. Application to a spiked oats sample is described.

Application of liquid chromatography‐atmospheric pressure chemical ionization mass spectrometry and tandem mass spectrometry to the determination of volatile nitrosamines in dry sausages

A high-performance liquid chromatographic-atmospheric pressure chemical ionization mass spectrometric method was developed for the determination of volatile nitrosamines in dry sausages. Tandem mass spectrometry was applied for the detection of N-nitrosopyrrolidine, N-nitrosodiethylamine and N-nitrosopiperidine. N-nitrosomethylamine was detected by using the selected ion monitoring mode. The occurrence of the four different nitrosamines was monitored in 27 dry sausage samples and a correlation was observed between N-nitrosopyrrolidine and biogenic amines. Nitroso compounds are thus not only formed in heated conditions but formation can also occur during ripening of dry sausages by reaction between residual nitrite and amines formed during the fermentation process.