Validation of Enzytec™ Liquid D-Lactic Acid for Enzymatic Determination of D-Lactic Acid in Selected Foods and Beverages: Official Method 2024.06 First Action.

BACKGROUND

D- and L-lactic acid are produced naturally by lactic acid bacteria and are found in fermented milk products, pickled vegetables, and cured meats. D-lactic acid is formed by some microorganisms only, e.g., Lactobacillus lactis and Leuconostoc cremoris. D-Lactic acid is not formed or only in traces by "higher organisms," e.g., by animals. Therefore, the presence of D-lactate may serve as an indicator for microbial contamination or spoilage, assuming that fermentation techniques have not been used.

OBJECTIVE

To validate the performance of the Enzytec™ Liquid D-Lactic acid for the determination of D-lactic acid in food and beverages such as milk and (fermented) milk products, fermented vegetable products, wines, beer, fruit and vegetable juices, and eggs and egg powder.

METHODS

The kit contains two ready-to-use components, which makes handling easy and suitable for automation. D-lactic acids react in the presence of NAD and D-lactate dehydrogenase to pyruvate and NADH. The NADH formed is equivalent to the amount of D-lactic acid converted.

RESULTS

Ascorbic acid, 3-hydroxybutyric acid, and sulfite interfere at concentrations higher than 0.2, 0.2, and 0.05 g/L, respectively. Oxaloacetic acid, pyruvic acid, and D-fructose do not interfere at or below concentrations of 0.2, 1, and 10 g/L, respectively. The calculated LOD when using a test volume of 100 µL is 5.4 mg/L, and the LOQ is 15 mg/L. The practical upper measurement range is 600 mg/L. Relative intermediate precision was between 3.5 and 5.7% for pineapple juice, sauerkraut juice, wine, and liquid egg. A reference material (wine) showed recoveries of 108%. For automation, three applications with different test volumes were validated. Linearity is given from 0.75 up to 3125 mg/L.

CONCLUSIONS

The method is robust and accurate and was approved as an AOAC Official Method of Analysis℠.

HIGHLIGHTS: The ready-to-use components of the test kit have a shelf life of at least 24 months.