Search: Ionenchromatographie Sugar alcohols

The sugar alcohols are separated using a strongly alkaline eluent and ion exchange column and detected and quantified electrochemically using a pulsed amperometric detector (PAD).

By applying a potential, the ions are oxidized at a gold electrode and induce a measurable charge. To prevent the electrode from being occupied in a very short time, the potential is then reversed to reduce and release the ions from the electrode.

Using a strongly alkaline eluent, carbohydrates are ionized and separated and quantified using an appropriate ion exchange column.

The carbohydrates are detected amperometrically. By applying a potential, the ions are oxidized at a gold electrode and induce a measurable charge. To prevent the electrode from being occupied in a very short time, the potential is then reversed to reduce and release the ions from the electrode.

The main components of wort are the fermentable sugars, monosaccharides, disaccharides and trisaccharides. Saccharomyces cerevisiae brewing yeast requires fermentable and assimilable sugars through oxidation, primarily glucose, fructose, sucrose, maltose and maltotriose, whereby wort is composed of predominantly maltose. Carbohydrates make up approximately 90 % of wort, 74 % of which is fermentable sugars. The total sugar concentration of a 12 %, standard-strength wort made from pale malt is 88 g/l on average.

The procedure for this analysis is analogous to method B-590.12.134 Sugar – HPLC or B-590.13.134 Fermentable carbohydrates – HPLC. In method B-590.12.134 Sugar – HPLC, an evaporative light scattering detector (ELSD) can be employed for the detection in addition to the refractometer (RI detector). If the ELSD is employed for detecting the sugar, aside from the isocratic elution, a gradient program can also be used. If the analysis is carried out using method B-590.13.134 Fermentable Carbohydrates – HPLC, one must ensure that the sample does not contain any maltose, since maltose and sucrose coelute.

Sucrose is important as a fermentable sugar for the technology of wort and beer production. Sucrose also plays a role in the evaluation and assessment of malt beverages and nutritional beers.

D-glucose content is determined before and after enzymatic hydrolysis of sucrose.

Sucrose is hydrolyzed by the enzyme β-fructosidase (invertase) at pH 4.6 to glucose and fructose:

\(\text{Sucrose + } H_2O \space {\xrightarrow{β-fructosidase}} \space \text{D-glucose + D-fructose}\)

Glucose is phosphorylated by the enzyme hexokinase (HK) and adenosine 5'-triphosphate (ATP) to glucose 6-phosphate (G-6-P):

\(\text{Glucose}+\text{ATP} \space \xrightarrow{HK} \space \text{G-6-P + ADP}\)

In the presence of the enzyme glucose-6-phosphate dehydrogenase (G6P-DH), G-6-P is oxidized from nicotinamide adenine dinucleotide phosphate (NADP) to gluconate-6-phosphate. Reduced nicotinamide adenine dinucleotide phosphate (NADPH) is formed:

\(\text{G-6-P + NADP} \hspace{0.8em} \xrightarrow{G6P-DH} \hspace{0.8em} \text{gluconate-6-phosphate + NADP + H}^+\)

The amount of NADPH formed during the reaction is equivalent to the amount of glucose. NADPH is a measurand and is determined on the basis of its absorbance at 334, 340 or 365 nm.

The sucrose content is calculated from the difference between the glucose concentration before and after enzymatic inversion.

D-Sorbit is present in pomaceous fruit and stone fruit.

D-Sorbitol is oxidized to D-fructose by nicotinamide adenine dinucleotide (NAD) in the presence of the enzyme sorbitol dehydrogenase (SDH), forming reduced nicotinamide adenine dinucleotide (NADH).

D-Sorbitol + NAD^{+ \(\xrightarrow{SDH}\)}D-G-6-P fructose + NADH + H⁺

The reaction equilibrium heavily favors the reactants NAD+ and D-sorbitol. It shifts toward D-fructose, when the NADH formed in this reaction reacts with iodine nitrotetrazolium chloride (INT) in a subsequent reaction. Formazan is created in the presence of diaphorase, which is an irreversible reaction.

NADH + INT + H⁺ \(\xrightarrow{diaphorase}\) NAD⁺ + Formazan

The maximum absorbance of the formazan is measured at 492 nm.

Separation by ion chromatography followed by conductivity detection.