Development of assays for coenzyme Q10 and vitamin K, and their application in clinical trials
Degree GrantorUniversity of Canterbury
Degree NameDoctor of Philosophy
This thesis describes the development of separate assays to measure coenzyme Q₁₀ (CoQ₁₀) and vitamin K. Coenzyme Q is essential for the mitochondrial electron transport chain, and vitamin K for the blood coagulation cascade. Vitamin K deficiency is associated with haemorrhagic disease of the new-born, and CoQ₁₀ deficiency with HMG-CoA-reductase inhibitor (statin) therapy and heart failure. Coenzyme Q and vitamin K are usually measured by HPLC, using electrochemical and ultraviolet, and electrochemical and fluorescence detection, respectively. For vitamin K1, the limit of detection achieved using fluorescence and electrochemical detection was 0.28 and 0.12 nmol/L, respectively. Sensitivity of fluorescence detection is improved by using protic solvents in the mobile phase, and platinum-black catalysed alcohol reduction. The lipophilicity and low endogenous concentrations of vitamin K1 hinder its measurement, and further work is required to produce a rapid, reliable and robust assay for its measurement in human plasma. The limits of detection achieved using fluorescence, ultraviolet and electrochemical detection to measure CoQ₁₀ were 29, 4.8, and 0.34 nmol/L, respectively. Plasma CoQ₁₀ is not stable during long term storage at -13 ℃, but at -80 ℃ it is stable for at least 18 months. The reference interval for plasma total CoQ₁₀ in the New Zealand population is 0.47 - 1.80 µmol/L. There is no clinical requirement for stratification of the reference interval according to gender. Coenzyme Q₁₀ in human plasma is homeostatically controlled, varying little over a two month interval in healthy young males. Coenzyme Q₁₀ supplements have significantly different bioavailability, with the median increase in plasma CoQ₁₀ ranging from 0.14 to 0.59 µmol/L for seven different supplement brands. There is a large inter-individual variation in CoQ₁₀ absorption, and hence plasma concentrations should be monitored during supplementation. A plateau in CoQ₁₀ absorption, from a single dose, at approximately 200 mg suggests that the maximum dose ingested at one time should be 200 mg or less. Q-Gel capsules containing 30 mg of CoQ₁₀ are twice as effective at raising blood CoQ₁₀ as 100 mg capsules. Plasma CoQ₁₀ in patients with chronic heart failure are significantly lowered by approximately 33% when these patients receive Atorvastatin for six weeks. The absolute decrease in CoQ₁₀ showed a significant correlation with worsening endothelial function (r = + 0.548, p = 0.011). Coenzyme Q₉ was shown to be present in human plasma with a reference interval of 8.8 - 47.0 nmol/L.