Der Ethanol-/Alkoholgehalt kann direkt mit den Methoden B-590.10.024 Stammwürze, Extrakt und Alkohol - destillativ (Amtliche Methode) und B-590.10.007 Stammwürze, Extrakt und Alkohol - Katalytische Verbrennung (SCABA) ermittelt werden.
Die Methoden B-590.10.070 Stammwürze, Extrakt und Alkohol - refraktiometrisch, B-590.10.026 Stammwürze, Extrakt und Alkohol - Biegeschwinger und Schallmessung, B-590.10.181 Stammwürze, Extrakt und Alkohol – Biegeschwinger und NIR und B-590.10.025 Stammwürze, Extrakt und Alkohol – thermoanalytisch ermitteln den Ethanol-/Alkoholgehalt indirekt.
Bei Bieren mit sehr geringem Ethanolgehalt kann aber die indirekte Messung ebenso wie die Destillationsanalyse Probleme hinsichtlich Richtigkeit der Ergebnisse bringen, so dass in solchen Fällen die enzymatische Analyse von Ethanol empfohlen wird [1, 2]. Die enzymatische Methode ist in die Amtliche Sammlung von Untersuchungsverfahren nach § 64 LFGB, 36.00/12 übernommen worden.
The concentrations of the coenzyme pairs NAD/NADH+ or NADP/NADPH+ are typically measured spectrophotometrically at a wavelength of 340 nm.
It is also possible to detect the coenzyme pairs photometrically at the wavelengths Hg 334 or Hg 365 nm.
Determination of ethanol by enzymatic methode (only alcohol dehydrogenase) for samples with low ethanol content.
Suitable for beers, non-alcoholic beers, reduced-alcohol beers, beer-based drinks, NAB, juice, beverages.
Ethanol is oxidized by nicotinamide adenine dinucleotide (NAD) in the presence of the enzyme alcohol dehydrogenase (ADH) to acetaldehyde:
\(\text{Ethanol + NAD}^+ \space ^{\underrightarrow{\text{ADH}}} \space \text{Acetaldehyd + NADH + H}^+\)
The equilibrium of this reaction favors the side with ethanol and NAD. In an alkaline medium, the equilibrium can be shifted to favor the substances on the right side of the equation.
The amount of NADH+H+ produced in the reaction is equivalent to the amount of ethanol and is measured photometrically due to its absorption at wavelengths of 340 nm.
Specificity of the determination [1]
The influence of aldehydes and ketones is eliminated by the order of reagent addition during the test. Methanol is not converted due to unfavorable KM values (Michaelis-Menten constant) of the enzymes used.
n-propanol and n-butanol are quantitatively converted under test conditions, higher primary alcohols lead to sample-dependent creep reactions. Secondary, tertiary and aromatic alcohols do not react. Glycerin does not interfere with the test even at higher concentrations.
Acetaldehyde does not interfere below a concentration of 3000 mg/l. Sulphite does not interfere below a concentration of 300 mg/l.