Rationale: Long-chain fatty acyl-coenzyme As (FA-CoAs) are important bioactive molecules, playing key roles in biosynthesis of fatty acids, membrane trafficking and signal transduction. Development of sensitive analytical methods for profiling theses lipid species in various tissues is critical to understand their biological activity. A high-pressure liquid chromatography/tandem mass spectrometry method has been developed for the quantitative analysis and screening of long-chain FACoAs in liver, brain, muscle and adipose tissue.
Methods: The sample preparation method consists of tissue homogenization, extraction with organic solvent and reconstitution in an ammonium hydroxide buffer. Extracts are separated by liquid chromatography (LC) on a reversed-phase column and detected by electrospray ionization tandem mass spectrometry (ESI-MS/MS) in positive mode. An additional neutral loss scan allows for untargeted FA-CoAs screening.
Results: Extraction was optimized for low sample load (10 mg) of four tissue types (liver, brain, muscle and adipose tissue) with recoveries between 60-140% depending on the analyte and tissue type. Targeted quantification was validated for ten FA-CoAs in the range 0.1-500 ng/mL with accuracies between 85-120%.
Conclusions: We have developed and validated a LC/MS/MS method for the quantifications and screening of long-chain FA-CoAs in four different types of mammalian tissue. The extraction method is straightforward and long-chain FA-CoA species can be quantified using only minimum amount of tissue. Copyright © 2016 John Wiley & Sons, Ltd.
Copyright © 2016 John Wiley & Sons, Ltd.