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  Section: Biotechnology Methods » Biochemistry
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Estimation of Sugar by Hagedorn-Jenson Method

The standard sodium is treated with potassium ferricyanide. A part of ferricyanide is reduced by glucose to ferrocyanide. The remaining ferricyanide is determined from the amount of iodine liberated. The ferrocyanide forms a salt with zinc and is not deoxidized to ferricyanide by atmospheric oxygen.
The principle reactions are

2K3 [Fe(CN)6] + 2KI →2K4 [Fe(CN)6] + I2

2K4 [Fe(CN)6] + ZnSO4 → K2Zn [Fe(CN)6] + 3K2SO4

The liberated iodine is estimated by titrating against sodium thiosulfate (0.005 N) using starch as an indicator. The amount of thiosulfate required is noted. The difference between the blank and the sample is measured. This shows the amount of thiosulfate required for sugar. By plotting a graph the concentration of sugar can be calculated.

Reagents Required

  1. Iodide sulfate chloride solution: 25 gms of ZnSO4 and 12.5 gms of NaCl2 are dissolved in 500 mL of water. To this 12.5 gms of KI is added on the day of the experiment.
  2. 3% acetic acid: 3 mL of acetic acid made up to 100 mL.
  3. Potassium ferricyanide solution K3 [Fe(CN)6]: 0.82 gm of K3 [Fe(CN)6] and 5.3 gms of anhydrous sodium carbonate is dissolved in water. Store it in a dark bottle.
  4. Preparation of standard sugar solution = 100 mg of sugar in 100 mL is prepared. Take 10 mL of stock and make it up to 100 mL.
  5. 0.1N Na2S2O3 solution: 12.4 gms of sodium thiosulfate in 500 mL of distilled water.
  6. Starch indicator: 1 gm of starch in 100 mL of H2O and 5 gms of NaCl.
  7. 0.1N K2Cr2O7: 0.49 gm in 100 mL of H2O.
Standardization of Na2S2O3 solution. Add 10 mL 0.1N K2Cr2O7 solution in a conical flask to 1 gm of KI and half of a test tube. H2SO4 1–2 drops of starch indicator and then titrate it against 0.1 N Na2S2O3 solution. The end point is blue to colorless. Seven clean and dry test tubes are used and sugar solutions ranging from 0.0, 0.4, 0.8, to 2 mL are added it is made up to 2 mL by adding distilled water. It produces 0–200 mg of sugar solution. Add 3 mL of potassium ferricyanide to each of the conical flasks. Boil the contents for 15 minutes and cool to room temperature. Then add 3 mL of iodine sulfate chloride solution and shake well. Add 2 mL of 3% acetic acid just before titrating against 0.005 N Na2S2O3 using the starch indicator. Titrate until the blue color disappears. Note the reading substrate from the first value. The values are plotted against the concentration of sugar solution from this graph. Concentration of unknown sugar is determined.

Result: The unknown solution contains ................ mg of sugar.


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