An Enzyme Fluorescence Quenching Method for the Determination of Lactate in Synthetic Blood Serum

An enzymatic fluorometric method for the determination of lactate based on its oxidation to pyruvate by NAD in the presence of lactate dehydrogenase (LDH) is proposed. The method relies on a decrease in the enzyme fluorescence during the reaction, which can be ascribed to the stability constant difference between the NAD-LDH and NADH-LDH complexes. A theoretical model was developed in order to predict the effect of the LDH and NAD concentrations on the analytical signal; also, the figures of merit of the proposed method were determined. The method allows the determination of lactate in synthetic blood serum with no interference, provided the sample is previously deproteinated with perchloric acid.