Bioluminescent determination of free fatty acids |
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Authors: | Horst Kather Eberhard Wieland |
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Institution: | Clinical Institute for Myocardial Infarction Research of the Medical Clinic of the University of Heidelberg, Bergheimerstraße 58, D-6900 Heidelberg, West Germany |
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Abstract: | A simple, highly specific, and sensitive bioluminescent method for determination of free fatty acids in unextracted plasma or serum has been developed. The method is based on the activation of free fatty acids by acyl-CoA synthetase (EC 6.2.1.3). The pyrophosphate formed is used to phosphorylate fructose 6-phosphate in a reaction catalyzed by the enzyme pyrophosphate-fructose-6-phosphate phosphotransferase (EC 4.1.2.13). The triosephosphates produced from fructose 1,6-bisphosphate by aldolase are oxidized by NAD in the presence of arsenate to 3-phosphoglycerate. The NADH is detected via the bacterial NADH-linked luciferase system. Excellent agreement has been obtained by comparison with accepted methods. In addition, for the determination of serum free fatty acids, the method is particularly applicable for following lipolysis of isolated adipocytes. |
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Keywords: | free fatty acids acyl-CoA synthetase NADH-linked luciferase system serum isolated rat adipocytes |
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