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Evaluation of soybean seed protein extraction focusing on metalloprotein analysis

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Abstract.

Two methods of protein extraction for soybean seeds were evaluated in terms of preservation of the metal ions bound to proteins after the extraction and separation procedures. The proteins were firstly separated according to their molar masses by polyacrylamide gel electrophoresis. Then, the protein bands were mapped by synchrotron radiation X-ray fluorescence in order to establish which metal ions were present in each one. Finally, some mapped protein bands were decomposed by microwave-assisted combustion and Ca, Cu, K, Mg, Mn, and Zn were quantified by inductively coupled plasma mass spectrometry or inductively coupled plasma optical emission spectrometry. The extraction methods studied were Method A (based on the treatment of ground soybean seeds with hexane and their extraction with Tris–HCl and β-mercaptoethanol) and Method B (based on the treatment of ground soybean seeds with petroleum ether and their extraction with Tris–HCl, dithiothreitol, phenylmethanesulfonyl fluoride, sodium dodecyl sulfate and potassium chloride). The best method was Method B, in which a 78% higher extraction efficiency was obtained when compared to Method A. Additionally, the metal-protein interactions were more appropriately preserved when Method B was applied, where the most affected ions were those that are bound weakly to proteins, such as Ca, K, and Mg.

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Authors and Affiliations

  1. Department of Analytical Chemistry, Institute of Chemistry, Universidade Estadual de Campinas (UNICAMP), PO Box 6154, 13084-971, Campinas, São Paulo, Brazil

    Alessandra Sussulini, Jerusa S. Garcia & Marco A. Z. Arruda

  2. Department of Chemistry, Universidade Federal de Santa Maria (UFSM), 97105-900, Santa Maria, Rio Grande do Sul, Brazil

    Márcia F. Mesko, Diogo P. Moraes & Érico M. M. Flores

  3. Laboratório Nacional de Luz Síncrotron (LNLS), PO Box 6192, 13084-971, Campinas, São Paulo, Brazil

    Carlos A. Pérez

Authors
  1. Alessandra Sussulini
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  2. Jerusa S. Garcia
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  3. Márcia F. Mesko
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  4. Diogo P. Moraes
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  5. Érico M. M. Flores
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  6. Carlos A. Pérez
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  7. Marco A. Z. Arruda
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Corresponding author

Correspondence to Marco A. Z. Arruda.

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Sussulini, A., Garcia, J., Mesko, M. et al. Evaluation of soybean seed protein extraction focusing on metalloprotein analysis. Microchim Acta 158, 173–180 (2007). https://doi.org/10.1007/s00604-006-0678-7

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  • DOI: https://doi.org/10.1007/s00604-006-0678-7

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