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

Posted on 21. May, 2012.

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How do fermentation and baking affect the structure, concentration and activity of chelating agents in home prepared bread? A study from the Czech Republic, reported in Chemical Speciation and Bioavailability, analyses alterations in the solubility and binding of trace elements that occur during the preparation of bread. Cereal grain proteins were found to be chelators of trace elements in the raw material. Some ligands of trace elements diminished during bread making, probably as the result of thermal denaturation.

ABSTRACT
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).

CONCLUSION
Our experiments considered some alterations in the solubility and binding of trace elements occurring during the preparation of bread. Using the IMAC technique and the analytical method of proteomics, we identified several cereal grain proteins as chelators of trace elements in the raw material for bread preparation. As most of these proteins are enzyme inhibitors, it seems that their biological role is not related to metal binding. Most of these proteins have a high content of cysteine or dicarboxylic amino-acids which causes an increased affinity for metal ions. However, this does not necessarily mean that these proteins are the original ligands of trace elements in cereal grains. Nevertheless, their participation in trace metal binding in flour, dough and bread is probable. The processes occurring in bread during fermentation and baking can affect the structure, concentration and activity of chelating agents.  The MALDI-MS analyses showed that some ligands of trace elements (namely the fraction of 12–14 kDa) are diminished during bread making, probably as the result of thermal denaturation.

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Keywords: bread, speciation, trace metals, mercury, proteomics

Doi: 10.3184/095422912X13325261626531

Oto Mestek, Jana Komínková, Jiří Šantrůček, Petr Kačer, Kateřina Mališová and Richard Koplík
Institute of Chemical Technology Prague, Technicka 5, 166 28 Prague 6, Czech Republic

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