Comparison of Two Enzyme Sequences for a Novel L-Malate Biosensor

Two novel amperometric biosensors for the determination of L -malic acid in food samples have been compared. Both sensors make use of a Clark-type O₂-electrode but differ in the enzymes used. The first sensor is composed of malate dehydrogenase (decarboxylating), also known as ‘malic enzyme’ (MDH(dec.), EC 1.1.1.40) and pyruvate oxidase (POP, EC 1.2.3.3). It covers a linear detection range from 1 μmol dm⁻³ to 0·9 mmol dm⁻³ L -malate, with a response time of 1·5 min (t₉₀) and a relative standard deviation of 3·5%. Measurements with real samples offered a good correlation with the standard enzymatic assay (difference ±7%). Stored at room temperature, the response of the sensor is constant for 8 days. The second biosensor is based on the three enzyme sequence malate dehydrogenase (MDH, EC 1.1.1.37), oxaloacetate decarboxylase (OAC, EC 4.1.1.3) and pyruvate oxidase (POP, EC 1.2.3.3). It has a non-linear calibration curve. Concentrations from 5 μmol dm⁻³ to 1 mmol dm⁻³ L -malate can be detected, within a response time of 1·5 min and with a relative standard deviation of 20%. The lower detection limit for L -malate is 2 μmol dm⁻³. The response is constant for 10 days when the sensor is stored at room temperature.