A Spectrophotometric Procedure for Malic Acid Determination in Wines Employing a Multicommutation Approach

This work describes an automated procedure to determine L-malic acid (MA) in wine samples using a multicommuted flow analysis. The MA quantification was based on an enzymatic reaction between MA and L-malate dehydrogenase (L-MDH) in the presence of nicotinamide adenine dinucleotide (NAD(+)), producing nicotinamide adenine dinucleotide dehydrogenase (NADH), which was monitored at 340 nm. The L-MDH was immobilized on a surface of modified silica with amino groups in the presence of glutaraldehyde. For studying optimization, the system was maintained with 200 mu L (288 U) of the L-MDH in 0.5 g of modified silica. Under the optimum experimental conditions, a linear response ranging from 0.1 to 1.5 g L-1 MA (R = 0.997 and n = 7), a detection (3 sigma criterion) and quantification (10 sigma criterion) limit estimated at 0.02 and 0.06 g L-1, respectively, a standard deviation relative of 1.8% (n = 7) for a sample of 0.5 g L-1 MA, a sampling rate of 67 samples per hour were achieved. Analyzing ten wines samples and applying the t-test to the results found and those obtained using reference procedures (HPLC) provided no significant differences at the 95% confidence level.