Analytical method development for selenium -containing proteins of clinical interest

2005 2005

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Abstract (summary)

The role of metallo-proteins in the body and their speciation is of increasing interest in clinical diagnostics because it is well recognized that their properties, function, and regulation in metabolic processes are highly species dependent. Therefore, their use as health markers for in vitro diagnostic, medical care and nutrition is directly related to their biological role. Selenium-containing proteins are important examples of speciation related health-indicators. In this work, significant progress has been made on the development of analytical methods for the speciation of selenoproteins in serum. A complete two-dimensional chromatographic separation of the different species of selenium-containing proteins of clinical interest including albumin, glutathione peroxidase and selenoprotein P is presented. Metal (Co 2+) chelate affinity high performance liquid chromatography (HPLC) was used to extract the proteins from serum, replacing the traditional low-pressure chromatography. Reversed phase chromatography was used for desalting and further separating selenoprotein fractions. The selenoproteins were identified by a combination of UV detection at 280 nm, selenium measurement by DRC-ICP-MS, and peptide mapping and molecular mass determination by MALDI-MS. Quantification of selenium in the selenium-containing protein species was accomplished using HPLC-UV-DRC-ICP-MS analysis employing 82Se stable isotope dilution. The sample was partially digested on line after separation, with nitric acid, using a novel in house designed interface (IRIS Digester-Evaporator, for which a patent has been applied). This interface reduces nitric acid and organic solvent from the liquid flow, and permits direct connection to the DRC-ICP-MS sample introduction system. It is a choice for HPLC-ICP-MS application or for faster on-line sample preparation for ICP and FAAS systems. The 2D chromatography, particularly the IMAC was the major source of variability in the overall method.

Indexing (details)

Analytical chemistry;
0486: Analytical chemistry
0487: Biochemistry
Identifier / keyword
Pure sciences; Cobalt ion chelate; Digester-evaporator; HPLC; Sample digestion; Selenium-containing proteins
Analytical method development for selenium -containing proteins of clinical interest
Arce-Osuna, Mariana
Number of pages
Publication year
Degree date
School code
DAI-B 66/10, Dissertation Abstracts International
Place of publication
Ann Arbor
Country of publication
United States
9780542382482, 0542382482
Uden, Peter C.
University of Massachusetts Amherst
University location
United States -- Massachusetts
Source type
Dissertations & Theses
Document type
Dissertation/thesis number
ProQuest document ID
Database copyright ProQuest LLC; ProQuest does not claim copyright in the individual underlying works.
Document URL
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