Enzyme-based amperometric biosensors for malic acid – A review

Highlights: • We summarise four decades of progress in developing amperometric malate biosensors. • We offer perspectives on the trends and strategies in malate biosensor design. • We compare and contrast the different enzymes that have been used over the years. • We highlight future directions, including bespoke enzymes and protein engineering. Malic acid is a key flavour component of many fruits and vegetables. There is significant interest in technologies for monitoring its concentration, particularly in winemaking. In this review we systematically and comprehensively chart progress in the development of enzyme-based amperometric biosensors for malic acid. We summarise the components and analytical parameters of malic acid sensors that have been reported over the past four decades, discussing their merits and pitfalls in terms of accuracy, sensitivity, linear range, response time and stability. We discuss how advances in electrode materials, electron mediators and the use of coupled enzymes have improved sensitivity and minimised interference, but also uncover a trade-off between sensitivity and linear range. A particular focus of our review is the three types of malate oxidoreductase enzyme that have been used in malic acid biosensors. We describe their different properties and conclude that identifying and/or engineering superior alternatives will be a key future direction for improving the commercial utility of malic acid biosensors.