12 Automatic Analysers in Oenology 655
Principle
This method enables the concentrations of glucose and fructose to be determined
separately or the sum of their concentrations to be determined in wines and musts.
Phosphorylation of glucose and fructose.
This reaction is catalysed by hexokinase (HK):HK
glucose+AT P←→glucose-6-phosphate+ADP (1)
HK
fructose+AT P←=>fructose-6-phosphate+ADPATP: adenosine triphosphate.
ADP: adenosine diphosphate.
Oxidation glucose-6-phosphate (G-6-P).
This reaction involves nicotinamide adenine dinucleotide phosphate (NADP), inthe presence of its specific enzyme, glucose-6-phosphate dehydrogenase (G6PDH).
G6PDH
G-6-P+NADP+←→gluconate-6-phosphate+NADPH+H+ (2)The equilibrium of the reaction is forcedin the direction of the products by the
appropriate of operating conditions (buffer pH 7.6 and excess NADP). The amount
of NADPH formed, which is proportional to the concentration of glucose in the
sample is measured by its absorbance at 340 nm.
This reaction, which enables the sum of glucose + fructose to be determined, is
catalysed by the enzyme phosphoglucose-isomerase (PGI):
PGI
fructose-6-phosphate←==>glucose-6-phosphate (3)The glucose-6-phosphate formed reacts with NADP as shown in reaction (2),
producing NADPH, which is measured by its absorbance at 340 nm.
The determination of fructose requires that the standards and samples are injected
twice, thus doubling the analysis time by comparison with the determination of
glucose alone.
12.2.2.7 Determination of Free and Total Sulphur Dioxide in Wines
and Musts by Dialysis
According to Scholten (1982) and Dubernet et al. (1997).
Principle
After acidification, free sulphur dioxide in the sample is allowed to diffuse across
a dialysis membrane. Combined sulphur dioxide is liberated by alkaline hydrolysis.