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In order to optimize the analytical method for the determination of fumonisin B1 (FB1) and fumonisin B2 (FB2) in different maize products, five materials (maize flour, corn flakes, extruded maize, muffins and infant formula) were investigated under a variety of experimental conditions organized in a ruggedness test according to a factorial design. The influence of five factors (extraction solvent, extraction mode, volume of extraction solvent, test sample size and clean-up) on method performances was tested by four laboratories using spiked materials (0.5 microgram/g and 1.5 micrograms/g FB1 + FB2) and naturally contaminated materials (ca 1.5 micrograms/g with FB1 + FB2). The end determination step was performed by high-performance liquid chromatography analysis of the o-phthaldialdehyde derivatized extracts. The ruggedness test permitted identification of two critical factors in the analysis of fumonisins in the above products, namely 'extraction solvent' and 'clean-up procedure'. In particular, the use of acetonitrile (ACN)-water (1 + 1, v + v) as extraction solvent and immunoaffinity column for clean-up provided better recovery of fumonisins and chromatographic resolution as compared with methanol (MeOH)-water (3 + 1, v + v) and strong anion exchange (SAX), respectively. However, phase separation occurring after extraction with ACN-water may have given incorrect results. Based on the information obtained with the present study it was possible to develop a new method horizontally applicable to all the above mentioned maize-based food matrices.

The determination of fumonisins in cornflakes is a challenging matter as the actually available methods for the analysis of corn do not perform well when applied to this more complex matrix. After testing several factors that may affect the analytical performance, an accurate method for the determination of fumonisin B1 (FB1) and B2 (FB2) in cornflakes has been developed. The method uses immunoaffinity chromatography for clean-up and high performance liquid chromatography (HPLC) for quantification of the toxins. Samples were extracted twice with acetonitrile-methanol-water (25:25:50) and the combined extracts were diluted with phosphate buffered saline (PBS) and applied to a FumoniTest immunoaffinity column. After washing with PBS, fumonisins were eluted from the column with methanol and reacted with o-phthaldialdehyde/2-mercaptoethanol to form fluorescent derivatives. Fumonisin derivatives were analysed by reversed phase HPLC with fluorometric detection using methanol-0.1 M phosphate buffer (77:23; pH adjusted at 3.35) as mobile phase. The average recoveries for FB1 and FB2 spiked in the ranges of 0.33 2.80 microg/g and 0.17-1.40 microg/g were 102.6% and 95.1%, respectively, with average relative standard deviations of 9% and 8%. The limit of quantification for FB1 and FB2 was 0.005 microg/g based on a signal-to-noise ratio of 6.1 by using a sensitive fluorescence detector. The method was used to analyse 18 cornflakes and cornflake cereals samples/for FB1 and FB2 contamination. All but one sample were found to be contaminated, with maximum FB1 and FB2 concentrations of 1.092 microg/g and 0.235 microg/g, respectively. Mean FB1 and FB2 concentrations were 0.157 microg/(g and 0.036 microg/g, respectively.

The effect of gamma-irradiation on fumonisin B1 (FB1) and fumonisin B2 (FB2) occurring in maize materials has been investigated together with the stability of fumonisins in gamma-irradiated maize stored at different temperatures (-18 to +40 degrees C) for different periods (2, 4, 13 and 26 weeks). Fifteen KGy gamma-irradiation was required to sterilize efficiently maize flour. This process caused a decrease in fumonisin content of about 20%. The stability studies showed that fumonisins are stable in gamma-irradiated maize for at least 6 months at 25 degrees C or at least 4 weeks at 40 degrees C. These data indicate that gamma-irradiation is an appropriate technique for obtaining sterilized maize materials to be used for intercomparison studies on analytical procedures for the measurement of the fumonisin content.