| Micro method for determination of nonesterified fatty acid in whole blood obtained by fingertip puncture. | |
| | |
MedLine Citation:
|
PMID: 16814738 Owner: NLM Status: MEDLINE |
Abstract/OtherAbstract:
|
Diagnostic tools for early identification of subjects at high risk for type 2 diabetes and other obesity-related disorders are important in prevention of these diseases. Nonesterified fatty acids (NEFAs) have been suggested to serve as a prediagnostic marker of diabetes and obesity-related disorders. In the current study, we developed a sensitive and reproducible micro method for quantification of NEFA in less than 10 microl whole blood. The method involves only two steps: (i) conversion of NEFA to fatty acid acyl-coenzyme A (acyl-CoA) esters using an acyl-CoA synthetase and (ii) quantification of the formed acyl-CoA esters with a fluorescent biosensor based on bovine acyl-CoA binding protein (ACBP). Lys50 of ACBP was mutagenized to a cysteine residue that was covalently modified with 6-bromoacetyl-2-dimethylaminonaphthalene to make a fluorescent acyl-CoA indicator (FACI-50). FACI-50 exhibits high fluorescence emission yield with maximum at 490 nm in the presence of CoA when excited at 387 nm. The addition of palmitoyl-CoA to a CoA-saturated FACI-50 lowered fluorescence emission by eightfold. Ethanol extract from 1 microl whole blood was incubated with ATP, CoA, and FACI-50. Following background fluorescence reading, NEFAs were converted to acyl-CoA by the acyl-CoA synthetase and the NEFA content was calculated from fluorescence emission changes using palmitic acid as external standard. The FACI-50 NEFA method was compared with two commercially available methods for quantification of NEFA. |
| | |
Authors:
|
Jesper S Hansen; Jens K Villadsen; Michael Gaster; Nils J Faergeman; Jens Knudsen |
Related Documents
:
|
9374128 - Phytanic acid alpha-oxidation: decarboxylation of 2-hydroxyphytanoyl-coa to pristanic a... 10359648 - Palmitoylation of the three isoforms of human endothelin-converting enzyme-1. 16657038 - Biosynthesis of fatty acids by a soluble extract from developing soybean cotyledons. 3174658 - Peroxisomal lignoceroyl-coa ligase deficiency in childhood adrenoleukodystrophy and adr... 4330148 - Ribosomal ribonucleic acid synthesis in bacillus subtilis. 3753138 - A sensitive, fast, durable, highly selective method for the determination of amino acid... |
Publication Detail:
|
Type: Journal Article; Research Support, Non-U.S. Gov't Date: 2006-05-19 |
Journal Detail:
|
Title: Analytical biochemistry Volume: 355 ISSN: 0003-2697 ISO Abbreviation: Anal. Biochem. Publication Date: 2006 Aug |
Date Detail:
|
Created Date: 2006-07-18 Completed Date: 2006-09-18 Revised Date: 2006-11-15 |
Medline Journal Info:
|
Nlm Unique ID: 0370535 Medline TA: Anal Biochem Country: United States |
Other Details:
|
Languages: eng Pagination: 29-38 Citation Subset: IM |
Affiliation:
|
Department of Biochemistry and Molecular Biology, University of Southern Denmark, DK-5230 Odense M, Denmark. |
Export Citation:
|
APA/MLA Format Download EndNote Download BibTex |
| MeSH Terms | |
Descriptor/Qualifier:
|
Acyl Coenzyme A
/
metabolism Animals Blood Glucose / metabolism Fatty Acid-Binding Proteins / chemistry, genetics, metabolism Fatty Acids, Nonesterified / analysis, blood*, standards Fingers / blood supply* Humans Lysine / chemistry, genetics, metabolism Mutagenesis, Site-Directed Mutant Proteins / chemistry, genetics, metabolism Palmitic Acid / analysis Palmitoyl Coenzyme A / analysis Protein Binding / genetics Protein Conformation Protein Structure, Tertiary Punctures / methods* Reference Standards Spectrometry, Fluorescence |
| Chemical | |
Reg. No./Substance:
|
0/Acyl Coenzyme A; 0/Blood Glucose; 0/Fatty Acid-Binding Proteins; 0/Fatty Acids, Nonesterified; 0/Mutant Proteins; 1763-10-6/Palmitoyl Coenzyme A; 56-87-1/Lysine; 57-10-3/Palmitic Acid |
From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine
Previous Document: Homogeneous time-resolved fluorescence and its applications for kinase assays in drug discovery.
Next Document: Control of laparoscopic instrument motion in an inanimate bench model: implications for the training...