Determination of citric acid by enzymatic means
This analysis is suitable for malt, wort, beer, beer-based beverages and soft drinks
Fruit juices:
The acid spectrum typical of certain types of fruit are used, along with other criteria, as a basis for recognizing unadulterated fruit juices. Tartaric acid, citric acid and L-malic acid are recorded here, which, with a few exceptions, determine the total acidity of the fruit.
Citric acid occurs as the primary acid in citrus juices and other juices. Orange juice usually contains 3–17 g/l citric acid (AIJN).
In citrus juices, an addition of citric acid can be detected via the citric acid/D-isocitric acid ratio, as this lies within relatively narrow limits. In orange juice, values below 130 are found.
D-isocitric acid is partly present in fruit products as a lactone. The lactone must first be saponified prior to enzymatic determination in order to detect the total D-isocitric acid content.
Malt, wort and beer:
Citric acid is an organic acid and is present in malt and wort and is also produced during fermentation.
Citric acid (citrate) is converted to oxaloacetic acid and acetic acid catalyzed by the enzyme citrate lyase (CL):
Citrate oxaloacetic \(^{\underrightarrow{CL}}\) acid + acetate
In the presence of the enzymes malate dehydrogenase (MDH) and lactate dehydrogenase (LDH), oxaloacetic acid and its decarboxylation product pyruvic acid are reduced to L-malic acid and L-lactic acid, respectively, by reduced nicotinamide adenine dinucleotide (NADH):
Oxaloacetate + NADH + H+ \(^{\underrightarrow{L-MDH}}\) L-malate + NAD+
Pyruvate + NADH + H+ \(^{\underrightarrow{L-LDH}}\)L-lactate + NAD+
The sum of the quantity of NADH consumed during the reaction is equivalent to the quantity of citric acid. The absorbance is determined photometrically at 334, 340 or 365 nm.
Mineral, spring and table waters as well as drinking waters that are to be marketed.
Qualitative detection of sulphite-reducing, spore-forming anaerobes in mineral water, spring water, table water, and other drinking water bottled for distribution to the consumer, by means of membrane filtration and immersion of the filter in single-concentrate dextrose-iron citrate-sodium sulphite broth (DRCM broth).
Note 1: If only the spores of sulphite-reducing anaerobes are to be analysed, the prescribed sample volume must be heated at 75 °C ± 1 °C for 10 minutes in order to inactivate the vegetative cells. Then cool quickly with cold water.
Important: Carry out a comparative test with the same volume of tap water to determine the total heating time required in the water bath.