ABSTRACT
In this study, the catalytic activity of aldehyde oxidase was investigated in various water-miscible organic solvents for the first time. The enzyme was partially purified from rabbit liver, and its activity was determined in the absence/presence of nine hydro-organic mixtures. The effects of pH and temperature on the enzyme activity were also investigated. The activity was reduced in the presence of all nine organic solvents. However, in some cases, the residual activity remained almost unchanged throughout the incubation of enzyme at 35 °C for 24 h. Considering potential advantages of doing reactions in the presence of organic solvents, these results could be of value. The enzyme showed different behavior in the reaction solutions making it difficult to formulate results in a single comprehensive model. The results indicated that binding and cleavage of the substrate are influenced in the presence of organic solvents.
Subject(s)
Aldehyde Oxidase/metabolism , Solvents/pharmacology , Water/chemistry , Enzyme Stability/drug effects , Hydrogen-Ion Concentration , TemperatureABSTRACT
Aldehyde oxidase is a widely distributed enzyme that is involved in the metabolism of an extensive range of aldehydes and N-heterocyclic compounds with physiological, pharmacological, and toxicological relevance. In the present study, a highly sensitive RP-HPLC-fluorescence method based on the oxidation of phenanthridine to phenanthridinone has been developed and validated to assay aldehyde oxidase activity in biological samples. Determination of phenanthridinone was achieved on a C18 column using 10 mmol/L phosphate buffer (pH 5.0) containing 0.1 mmol/L EDTA-acetonitrile (40 + 60, v/v) as the mobile phase. The fluorescence intensity of phenanthridinone was measured at 364 nm with excitation at 236 nm. The proposed method was precise, accurate, specific and rapid (analysis time, approximately 8 min) with a mean RSD of 2.54%. Peak responses were linear from 0.5 to 100 nmol/L, with an LOD of 0.125 nmol/L. The applicability of the method was demonstrated by measurement of aldehyde oxidase activity in rat liver, kidney, ovary, and heart fractions.
