Determination of coenzyme Q biosynthesis in cultured cells without the necessity for lipid extraction

Ubiquinone (coenzyme Q; CoQ) is the only lipophilic antioxidant that is endogenously synthesized by all organisms. CoQ biosynthesis is determined in vitro by supplying a radiolabeled precursor and, after lipid extraction and CoQ separation by thin-layer chromatography or high-performance liquid chro...

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Bibliographic Details
Published in:Analytical biochemistry 2005, Vol.336 (1), p.60-63
Main Authors: Córdoba-Pedregosa, María del Carmen, Villalba, José M., Alcaín, Francisco J.
Format: Article
Language:English
Subjects:
Carbon Radioisotopes
Carcinoma, Hepatocellular - metabolism
Humans
Parabens - metabolism
Tumor Cells, Cultured - metabolism
Ubiquinone - analysis
Ubiquinone - biosynthesis
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Summary:Ubiquinone (coenzyme Q; CoQ) is the only lipophilic antioxidant that is endogenously synthesized by all organisms. CoQ biosynthesis is determined in vitro by supplying a radiolabeled precursor and, after lipid extraction and CoQ separation by thin-layer chromatography or high-performance liquid chromatography, the radioactivity present in the sample is quantified. In the rapid and simple method described here, we avoid the use of organic solvents by supplying 4-hydroxy-[U- 14C]benzoate as radiolabeled precursor and precipitating CoQ with trichloroacetic acid (TCA). After TCA precipitation, all radioactivity was present in the precipitate and CoQ was the only radiolabeled molecule detected. The radioactive material was then solubilized with NaOH and quantified in a scintillation counter.
ISSN:0003-2697
1096-0309
DOI:10.1016/j.ab.2004.09.031