Analyses of trace metals, peptide ligands of trace metals and mercury speciation in home prepared bread

Bread represents an important source of trace elements in the human diet. This study is focused on home prepared bread in the Czech Republic. The amounts of Cu, Mo, Mn, Ni and Zn (total and soluble in Tris-HCl buffer, pH 7.5), Cd, Co, Fe, Pb and Tl (total only) as well as Hg (total and soluble in mercaptoethanol-HCl mixture) in raw materials and baked bread were determined using ICP-MS. Moreover, the speciation of elements was investigated using HPLC/ICP-MS. Isolated peptide ligands of the trace elements were analysed for amino acids and characterised by MALDI-MS. The concentrations of all elements were in accordance with Czech legislation. The solubility of the Ni species was not affected by the baking process, whereas the solubilities of Mo, Mn and Zn species decreased. Soluble mercury was found only in the inorganic form. The soluble species of Cu, Mo, Mn, Ni and Zn were found in two fractions with the apparent molecular weights of 1-2 kDa and 4-5 kDa. Ligands of trace metals isolated from these fractions contained appreciable amounts of Asx, Glx, Gly, Ser and Cys. No phytochelatin-like peptides were found in the MALDI-MS spectra of isolated ligands,. Using MALDI-MS/MS, the partial amino-acids sequences of peptide ligands were obtained, and the linkages of peptides and saccharides confirmed. The MS analysis of the trypsin digest of the medium molecular weight fraction revealed several proteins rich in cysteine (e.g., barwin and amylase inhibitors).