Measurement of reduced and oxidized coenzyme Q9 and coenzyme Q10 levels in mouse tissues by HPLC with coulometric detection

Abstract Background: Ubiquinone-responsive multiple respiratory chain dysfunction due to coenzyme Q10 (CoQ10) deficiency has been previously identified in muscle biopsies. However, previous methods are unreliable for estimating CoQ10 redox status in tissue. We developed an accurate method for measuring tissue concentrations of reduced and oxidized coenzyme Q (CoQ). Methods: Mouse tissues were weighed in the frozen state and homogenized with cold 1-propanol on ice. After solvent extraction, centrifugation and filtration, the filtrate was subsequently analyzed by reversed-phase HPLC with coulometric detection. Results: Reference calibration curves were used to determine reduced and oxidized coenzyme Q9 (CoQ9) and CoQ10 concentrations in tissues. The method is sensitive (∼15 μg/l), reproducible (6% CV) for CoQ9 and CoQ10, and linear up to 20 mg/l for CoQ9 and CoQ10. Analytical recoveries were 90–104%. In mouse tissues the amounts of total CoQ (TQ) ranged from 261 to 1737 nmol/g of protein. Total CoQ9 levels are comparable with the values of those previously reported. CoQ is found to be mostly in the reduced form in mouse liver (∼87%), heart (∼60%), and muscle tissues (∼58%); in the brain, most of the CoQ is in the oxidized state (∼65%). Conclusion: This procedure provides a precise, sensitive, and direct assay method for the determination of reduced and oxidized CoQ9 and CoQ10 in mouse hindleg muscle, heart, brain, and liver tissues.