Metallomics Approach to Trace Element Analysis in Ustilago maydis Using Cellular Fractionation, Atomic Absorption Spectrometry, and Size Exclusion Chromatography with ICP-MS Detection

Abstract

Huitlacoche is the ethnic name of the young fruiting bodies of Ustilago maydis, a common parasite of maize. In Mexico and other Latin American countries, this fungus has been traditionally appreciated as a local delicacy. In this work a metallomics approach was used with the determination of eight elements in huitlacoche by electrothermal atomic absorption spectrometry as one facet of this approach. The results obtained indicated relatively lower concentrations of commonly analyzed metals, as referred to the data reported for other mushroom types. This effect was ascribed to different accessibilities of elements, depending on fungus substrate (lower from plant than from soil). Subcellular fractionation was accomplished by centrifugation of cell homogenates suspended in Tris-HCl buffer. Recoveries of the fractionation procedure were in the range of 71-103%. For six elements (Cr, Cu, Fe, Mn, Ni, and Pb), the mean relative contributions in cytosol, cell walls, and mixed membrane fraction were 50.7, 48.2, and 1.1% respectively. To attain the molecular weight distribution of compounds containing target elements as an additional aspect of the metallomics approach, the fungus extract (1% sodium dodecyl sulfate in Tris-HCl, 30 mmol L(-)(1), pH 7.0) was analyzed by size exclusion chromatography with UV and ICP-MS detection. With spectrophotometric detection (280 nm), the elution of high molecular weight compounds was observed in the form of one peak (MW > 10 kDa), and several lower peaks appeared at higher retention times (MW < 10 kDa). On ICP-MS chromatograms, a coelution of (59)Co, (63)Cu, (57)Fe, (202)Hg, (60)Ni, and (80)Se with the first peak on the UV chromatogram was clearly observed, indicating that a fraction of each element incorporated with high molecular weight compounds (12.7, 19.8, 33.7, 100, 19.4, and 45.8%, respectively, based on the peak area measurements). From a comparison of (80)Se and (33)S chromatograms (for sulfur analysis, the extract was obtained in the absence of SDS), both elements coeluted with the first UV peak, but their lower molecular weight compounds were apparently different. These findings may contribute to a better understanding of the accumulation of elements in mushrooms.