HPLC analysis of reduced and oxidized coenzyme Q(10) in human plasma.

BACKGROUND The percentage of reduced coenzyme Q(10) (CoQ(10)H(2)) in total coenzyme Q(10) (TQ(10)) is decreased in plasma of patients with prematurity, hyperlipidemia, and liver disease. CoQ(10)H(2) is, however, easily oxidized and difficult to measure, and therefore reliable quantification of plasma CoQ(10)H(2) 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 degrees C. The plasma was harvested and stored in screw-top polypropylene tubes at -80 degrees 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 CoQ(10) into CoQ(10)H(2) and avoided artifactual oxidation of CoQ(10)H(2). The electrochemical reduction yielded 99% CoQ(10)H(2). Only 100 microL of plasma was required to simultaneously measure CoQ(10)H(2) and CoQ(10) over an analytical range of 10 microg/L to 4 mg/L. Intra- and interassay CVs for CoQ(10) in human plasma were 1.2-4.9% across this range. Analytical recoveries were 95.8-101.0%. The percentage of CoQ(10)H(2) in TQ(10) was approximately 96% in apparently healthy individuals. The method allowed analysis of up to 40 samples within an 8-h period. CONCLUSIONS This optimized method for CoQ(10)H(2) 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.