Determination of xanthohumol and isoxanthohumol
All beers, beer-based beverages, wort, ethanol extracts, CO2 spent hops and xanthohumol products
Xanthohumol and isoxanthohumol are dissolved with acetonitrile from the sample and following separation, are determined using a Nucleodur C18 column and UV detection.
The method is suitable for beers of all original wort ranges and alcohol contents.
Volatile compounds in beer are concentrated through distillation and the distillate is quantitatively determined by direct injection into a gas chromatograph. The linearity of the detector and the determination of the concentrations of analytes in the sample are achieved by using multiple concentration levels within the relevant range and through evaluation of the relative area under the peaks.
This method is suitable for beers exhibiting all ranges of original gravity and alcohol concentrations.
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.
A known quantity of an internal standard (n-propanol) is added to the sample, if necessary after dilution.
The sample is equilibrated at a certain temperature in a headspace vial and part of the headspace is injected into a gas chromatograph.
The ethanol contained is separated on a polar gas chromatography column and detected with a flame ionization detector (FID).
The ethanol concentration in % vol. is calculated from the ratio of the area of the ethanol peak to the area of the internal standard (n-propanol) with the ratios of the same peaks determined when analyzing standards with known ethanol concentrations.
The method is suitable for the determination of water vapor volatile aroma compounds in beer.
Volatile aroma compounds are driven out of the sample through steam distillation. The ethanolic distillate is saturated with NaCl. Potassium hydrogen sulfite is added to separate carbonyl groups that might interfere with the analysis. The extraction of the aroma compounds is performed by shaking out with dichloromethane and the phases separated by centrifuging.
Determination of ethanol by enzymatic method with two enzymes (alcohol dehydrogenase and aldehyde 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 and through removal of the acetaldehyde produced, the equilibrium can be shifted to favor the substances on the right side of the equation. Acetaldehyde is quantitatively oxidized to acetic acid in the presence of aldehyde dehydrogenase (Al-DH):
\(\text{Acetaldehyde + NAD}^+ \space ^{\underrightarrow{\text{Al-DH}}} \space \text{acetic acid + NADH + H}^+\)
The amount of NADH+H+ produced in the reaction is equivalent to half of 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.