Search: Glucose

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

Glucose + ATP \(^{\underrightarrow{HK}}\) G-6-P + ADP

Fructose + ATP \(^{\underrightarrow{HK}}\) F-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:

\(\text{G-6-P}\hspace{0.2em}+\hspace{0.2em}\text{NADP}\hspace{0.8em}^{\underrightarrow{\text{G6P–DH}}}\hspace{0.8em} \text{glucanate-6-phosphate} + \text{NADP}+\text{H}^+\)

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

After the reaction is complete, F-6-P is converted to G-6-P by phosphoglucose isomerase (PGI):

F-6-P \(^{\underrightarrow{PGI}}\) G-6-P

G-6-P reacts in turn with NADP to form gluconate-6-phosphate and NADPH. The additional amount of NADPH formed is equivalent to the amount of fructose and is determined photometrically based on its absorption at 334, 340 or 365 nm.

The D-glucose content is determined before and after enzymatic hydrolysis of sucrose. D-fructose is measured following D-glucose determination.

D-glucose determination before inversion:

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

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

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:

\(\text{G-6-P + NADP} \space ^{\underrightarrow{\text{G6P-DH}}} \space \text{gluconate-6-phosphate + NADP + H}^+\)

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

D-fructose determination:

Hexokinase catalyzes the phosphorylation of D-fructose with ATP to D-fructose-6-phosphate.

\(\text{Fructose + ATP} \space ^{\underrightarrow{\text{HK}}} \space \text{F-6-P + ADP}\)

After the reaction is complete, F-6-P is converted to G-6-P by phosphoglucose isomerase (PGI):

\(\text{F-6-P} \space ^{\underrightarrow{\text{PGI}}} \space \text{G-6-P}\)

G-6-P reacts in turn with NADP to form gluconate-6-phosphate and NADPH. The additional amount of NADPH formed is equivalent to the amount of fructose and is determined photometrically based on its absorbance at 334, 340 or 365 nm.

Enzymatic inversion:

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

\(\text{Sucrose}+H_2O\hspace{0.8em} ^{\underrightarrow{\text{B-fructosidase}}} \hspace{0.8em} \text{glucose + fructose}\)

The D-glucose determination after inversion (total D-glucose) is carried out as described above.

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

Manganese ions are catalytically oxidized into purple-colored permanganate ions. The purple color of the solution is determined using a comparator (or photometrically):

Mn²⁺ + 12  H₂O → MnO₄^- + 8 H₃O⁺ + 5 e^-

Potassium permanganate oxidizes many organic and certain inorganic substances more or less completely in acidic, neutral or alkaline solutions. The volume of potassium permanganate required in the analysis is determined potentiometrically. Since oxidation depends on the type of solution, on its temperature and on the reaction time, the procedure described below must be followed precisely.

In acidic solutions, permanganate ions are typically reduced to manganese(II) ions:

MnO₄^- + 5 e- + 8 H₃O⁺ → Mn²⁺ + 12 H₂O

In alkaline solutions, the reduction results in tetravalent manganese only:

MnO₄^- + 3 e- + 4 H₃O⁺ → MnO₂ + 6 H₂O

Since in both cases the titration takes place in an acidic solution, this is irrelevant for the calculation. By adding oxalic acid, both the excess permanganate ions as well as the tetravalent manganese are reduced to manganese(II) ions:

2 MnO₄^- + 5 C₂O₄^2- + 16 H₃O⁺ → 2 Mn²⁺ + 24 H₂O + 10 CO₂

MnO₂ + C₂O₄²- + 4 H₃O⁺ → Mn²⁺ + 6 H₂O + 2 CO₂  

Carbohydrates are split by acid hydrolysis in the boiling heat under reflux with acid. After neutralization and filtration of the hydrolysis mixture, the glucose content is determined enzymatically after dilution. In this process, glucose is phosphorylated by the enzyme hexokinase (HK) and adenosine 5'-triphosphate (ATP) to glucose 6-phosphate (G-6-P):

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

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

\(\text{G-6-P + NADP}^+ \space ^{\underrightarrow{\text{G6P-DH}}} \space \text{NADPH + H}^+\)

The amount of NADPH formed during the reaction is equivalent to the amount of glucose. NADPH is determined based upon its absorbance at 334, 340 or 365 nm.

The quantitative determination is carried out in a sulphuric acid solution by titration with potassium permanganate:

5 H₂0₂ +  2  KMnO₄ + 3  H₂S0₄  →  2 MnSO₄ + H₂S0₄ + 8 H₂0 + 5 0₂

If the sample contains other peroxide-containing disinfectants in addition to hydrogen peroxide (H₂O₂), the method T-754-01-032 Peracetic-acid-based-disinfectants must be used.