Method to assess fatty acid ethyl ester binding to albumin

Catherine A. Best, Michael Laposata, Veronia G. Poios, Zbigniew M. Szczepiorkowski

Research output: Contribution to journalArticle

10 Citations (Scopus)

Abstract

Aims: To develop a method to assess the relative binding of fatty acid ethyl esters (FAEE) and free fatty acids (FFA) to albumin, and to determine if binding affinity is related to fatty acid chain length and or degree of saturation. Methods: Radiolabelled ethyl-[14C] oleate-b ound to albumin was challenged with various ratios of FFA to FAEE. The displacement of ethyl-[ 14 C] oleate by FFA was visualized and quantitated through a combination of native-PAGE, autoradiography, and liquid scintillation counting (LSC). Results: As the ratio of FFA to FAEE increased from 0:1 to 12:1, for all fatty acids tested (myristate, palmitate, stearate, oleate, linoleate, and arachidonate), ethyl-[14C] oleate displacement increased, when expressed as radioactivity (in DPM) as a percentage of control. In contrast, ethyl oleate did not displace stearate or oleate from albumin at molar ratios up to 5:1 (FAEE:FFA). Conclusions: The method developed gave reproducible visualization of noncovalent binding of radiolabelled FAEE to albumin. The combination of native-PAGE and autoradiography LSC works well in assessing the binding properties of albumin and radiolabelled FAEE. The data indicate preferential binding of FFA over FAEE to albumin with six different FFA displacing FAEE to an approximately equal extent.

Original languageEnglish (US)
Pages (from-to)240-246
Number of pages7
JournalAlcohol and Alcoholism
Volume41
Issue number3
DOIs
StatePublished - May 2006
Externally publishedYes

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Albumins
Esters
Fatty Acids
Nonesterified Fatty Acids
Oleic Acid
Native Polyacrylamide Gel Electrophoresis
Stearates
Scintillation Counting
Scintillation
Autoradiography
Palmitates
Radioactivity
Myristic Acid
Liquids
Linoleic Acid
Chain length
Visualization
ethyl oleate

ASJC Scopus subject areas

  • Medicine (miscellaneous)
  • Toxicology

Cite this

Method to assess fatty acid ethyl ester binding to albumin. / Best, Catherine A.; Laposata, Michael; Poios, Veronia G.; Szczepiorkowski, Zbigniew M.

In: Alcohol and Alcoholism, Vol. 41, No. 3, 05.2006, p. 240-246.

Research output: Contribution to journalArticle

Best, Catherine A. ; Laposata, Michael ; Poios, Veronia G. ; Szczepiorkowski, Zbigniew M. / Method to assess fatty acid ethyl ester binding to albumin. In: Alcohol and Alcoholism. 2006 ; Vol. 41, No. 3. pp. 240-246.
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N2 - Aims: To develop a method to assess the relative binding of fatty acid ethyl esters (FAEE) and free fatty acids (FFA) to albumin, and to determine if binding affinity is related to fatty acid chain length and or degree of saturation. Methods: Radiolabelled ethyl-[14C] oleate-b ound to albumin was challenged with various ratios of FFA to FAEE. The displacement of ethyl-[ 14 C] oleate by FFA was visualized and quantitated through a combination of native-PAGE, autoradiography, and liquid scintillation counting (LSC). Results: As the ratio of FFA to FAEE increased from 0:1 to 12:1, for all fatty acids tested (myristate, palmitate, stearate, oleate, linoleate, and arachidonate), ethyl-[14C] oleate displacement increased, when expressed as radioactivity (in DPM) as a percentage of control. In contrast, ethyl oleate did not displace stearate or oleate from albumin at molar ratios up to 5:1 (FAEE:FFA). Conclusions: The method developed gave reproducible visualization of noncovalent binding of radiolabelled FAEE to albumin. The combination of native-PAGE and autoradiography LSC works well in assessing the binding properties of albumin and radiolabelled FAEE. The data indicate preferential binding of FFA over FAEE to albumin with six different FFA displacing FAEE to an approximately equal extent.

AB - Aims: To develop a method to assess the relative binding of fatty acid ethyl esters (FAEE) and free fatty acids (FFA) to albumin, and to determine if binding affinity is related to fatty acid chain length and or degree of saturation. Methods: Radiolabelled ethyl-[14C] oleate-b ound to albumin was challenged with various ratios of FFA to FAEE. The displacement of ethyl-[ 14 C] oleate by FFA was visualized and quantitated through a combination of native-PAGE, autoradiography, and liquid scintillation counting (LSC). Results: As the ratio of FFA to FAEE increased from 0:1 to 12:1, for all fatty acids tested (myristate, palmitate, stearate, oleate, linoleate, and arachidonate), ethyl-[14C] oleate displacement increased, when expressed as radioactivity (in DPM) as a percentage of control. In contrast, ethyl oleate did not displace stearate or oleate from albumin at molar ratios up to 5:1 (FAEE:FFA). Conclusions: The method developed gave reproducible visualization of noncovalent binding of radiolabelled FAEE to albumin. The combination of native-PAGE and autoradiography LSC works well in assessing the binding properties of albumin and radiolabelled FAEE. The data indicate preferential binding of FFA over FAEE to albumin with six different FFA displacing FAEE to an approximately equal extent.

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