HPLC Analysis of Reduced and Oxidized Coenzyme Q10 in Human Plasma

Abstract

AbstractBackground: The percentage of reduced coenzyme Q10 (CoQ10H2) in total coenzyme Q10 (TQ10) is decreased in plasma of patients with prematurity, hyperlipidemia, and liver disease. CoQ10H2 is, however, easily oxidized and difficult to measure, and therefore reliable quantification of plasma CoQ10H2 is of clinical importance.Methods: Venous blood was collected into evacuated tubes containing heparin, which were immediately placed on ice and promptly centrifuged at 4 °C. The plasma was harvested and stored in screw-top polypropylene tubes at −80 °C until analysis. After extraction with 1-propanol and centrifugation, the supernatant was injected directly into an HPLC system with coulometric detection.Results: The in-line reduction procedure permitted transformation of CoQ10 into CoQ10H2 and avoided artifactual oxidation of CoQ10H2. The electrochemical reduction yielded 99% CoQ10H2. Only 100 μL of plasma was required to simultaneously measure CoQ10H2 and CoQ10 over an analytical range of 10 μg/L to 4 mg/L. Intra- and interassay CVs for CoQ10 in human plasma were 1.2–4.9% across this range. Analytical recoveries were 95.8–101.0%. The percentage of CoQ10H2 in TQ10 was ∼96% in apparently healthy individuals. The method allowed analysis of up to 40 samples within an 8-h period.Conclusions: This optimized method for CoQ10H2 analysis provides rapid and precise results with the potential for high throughput. This method is specific and sufficiently sensitive for use in both clinical and research laboratories.