Validation of Enzytec™ Liquid D-Gluconic Acid for Enzymatic Determination of D-Gluconic Acid and Glucono-δ-lactone in Selected Foods and Beverages: First Action 2025.05

Background D-gluconic acid is found in honey, ripe fruits and wine. In food industry, D-gluconic acid is used as a complexing agent and acidifier in desserts, beverages, fruit and vegetable products. D-glucono-δ-lactone is derived from gluconic acid, which hydrolyzes slowly with the release of protons. It can be used wherever slow acidification is desired. Objective To validate the Enzytec™ Liquid D-Gluconic acid test kit for the determination of the sum of D-gluconic and D-glucono-δ-lactone in food and beverages such as fruit juices, wine, fermented soft drinks, kombucha, and heated sausage meat. Methods The kit contains two ready-to-use components which makes handling easy and suitable for automation. Gluconic acid reacts with ATP in the presence of gluconate kinase to D-gluconate-6-phosphate and ADP followed by an indicator reaction that produces NADH, which is measured at 340 nm. Results The test is specific to D-gluconic acid and shows no side activities or interferences to relevant acids at or below 6.55 g/L. SO2 does not interfere at or below 0.5 g/L. Linearity for a test volume of 100 µL is given from 6 to 1500 mg/L. For 1000 µL, the LOQ is 0.8 mg/L. Intermediate precision is between 2.73 and 7.2% for different matrixes. Mean recovery for an incurred meat sausage is 84% at 799 mg/kg and 96% for the higher level. Spiking of the other matrixes resulted in recoveries between 96 and 103%. For automation, three applications with different test volumes were validated. Linearity is given from 2 up to 9375 mg/L. Conclusions The method is robust and accurate for manual and automated applications. The method was approved as AOAC Official Method of Analysis℠. Highlights The components of the test kit have a shelf life of at least 24 months.