Miniaturized HPLC-UV method for clinical Q10 analysis and its application in the treatment of mithocondrial disease with different Q10 formulations

CONTIN M.,; FLOR S.,; LUCANGIOLI S.,; TRIPODI V.

Belgica

Simposio; The 6th Conference of the international Coenzyme Q10 Assosiation; 2010

Coenzyme Q10 Assosiation

The method currently employed for determination of Coenzyme Q10 (Q10) in plasma or biological tissues is liquid chromatography with electrochemical detection (ECD) and it is chosen for its high selectivity and sensitivity. However, a long time is necessary to obtain a stabilized baseline before analysis, and due to the fact that ECD is sensitive to high concentrations of some lipophilic components that could be present in the sample, the electrodes could be passivated and reduced their lifetime. To achieve rapid analysis with high sensitivity, miniaturization of the column is a good alternative to the traditional chromatographic method. The use of a short column with reduced diameter allows the quantitation of Q10 in a shorter time with less consumption of solvents and reduced sample volumes. In this work we have developed a simple, rapid and highly sensitive method for routine analysis of Q10, specially apt for clinical laboratories using a miniaturized HPLC-UV method and more simple by the employement of UV detection. An analytical column of XTerra (50 mm x 2.1 mm i.d., 3.5 µm) filled with special hybrid particles was used. Chromatographic conditions were: 30ºC column temperature, 1 µl injection volume at 275 nm UV detection using a mobile phase consisting of 98% methanol and 2% water. For quantitation of Q10 in human plasma the sample was treated by 1-propanol precipitation followed by hexane extraction. Time of analysis was 5 minutes when a flow rate of 0.3 mL min -1 was applied. The developed method was found to be linear, precise and accurate. The LOD and LOQ of Q10 in plasma were 19 pg and 63 pg on column respectively, resulting to be comparable to the results of conventional HPLC methods with ECD. This method demonstrated to be useful to evaluate Q10 plasma concentration in a patient with mithocondrial disease during treatment with Q10 capsules formulated at the same dose but from different laboratories. We observed a reduction in plasma Q10 levels together with the reappearance of the symptoms in the patient treated with a Q10 formulation containing polyvinyl pirrolydone (PVP) in the excipients. It is important to emphasize that a continuous control of the plasma Q10 concentrations together with the quality control of the formulation administered are necessary in the treatment of associated diseases.