Synthesis and Characterization of Pyridine Compounds for Amperometric Measurements of Leukocyte Esterase

Abstract

Abstract We introduce a new class of substrates (compounds I – III ) for leukocyte esterase (LE) that react with LE yielding anodic current in direct proportion to LE activity. The kinetic constants K m and k cat for the enzymatic reactions were determined by amperometry at a glassy carbon electrode. The binding affinity of I – III for LE was two orders of magnitude better than that of existing optical LE substrates. The specificity constant k cat / K m was equal to 2.7, 3.8, and 5.8×10 5 m −1 s −1 for compounds containing the pyridine ( I ), methoxypyridine ( II ), and (methoxycarbonyl)pyridine ( III ), respectively, thus showing an increase in catalytic efficiency in this order. Compound III had the lowest octanol/water partition coefficient (log p =0.33) along with the highest topological surface area (tPSA=222 Å 2 ) and the best aqueous solubility (4.0 mg mL −1 ). The average enzymatic activity of LE released from a single leukocyte was equal to 4.5 nU when measured with compound III .