Reagents for Glucose Determination
24 Questions
1 Views

Reagents for Glucose Determination

Created by
@FriendlyTrust

Podcast Beta

Play an AI-generated podcast conversation about this lesson

Questions and Answers

How long can glucose oxidase reagent maintain its activity if refrigerated?

  • Indefinitely
  • Only 1 week
  • At least 2 weeks (correct)
  • Several months

    • What must be done to the concentration of glucose after preparing a protein-free supernatant from serum?

  • Multiply the answer by 25 (correct)
  • Calculate directly from the test result
  • Multiply the answer by 50
  • No calculation is necessary

    • Which reagent is used for protein precipitation and removal from serum?

  • Copper Sulphate Solution
  • Copper Sulphate solution
  • Sodium Tungstate solution (correct)
  • Isotonic Sodium Sulphate (correct)

    • What is the role of Cu2+ in the determination of blood glucose?

    <p>It is reduced to Cu+ during the reaction.</p> Signup and view all the answers

    What is the function of 1N-H2SO4 in the blood glucose determination method?

    <p>To stop enzyme action and stabilize the color</p> Signup and view all the answers

    What is the stability condition of standard glucose solutions at room temperature?

    <p>Stable indefinitely</p> Signup and view all the answers

    What does the formation of molybdenum blue indicate in colorimetric analysis?

    <p>Presence of reducing sugars.</p> Signup and view all the answers

    What is the purpose of adding 0.5 ml of glucose oxidase in the preparation of the glucose oxidase reagent?

    <p>To enhance the reaction with reducing sugars</p> Signup and view all the answers

    At what wavelength is colorimetric analysis performed in glucose detection?

    <p>650 nm</p> Signup and view all the answers

    What is the purpose of adding Na2SO4 in the process described?

    <p>To prevent oxidation of Cu2O.</p> Signup and view all the answers

    How is the acetate buffer prepared for use in the glucose oxidase reagent?

    <p>Add 3 volumes of sodium acetate to 7 volumes of acetic acid</p> Signup and view all the answers

    What is the purpose of protein precipitation in the glucose determination method?

    <p>To remove proteins that interfere with analysis</p> Signup and view all the answers

    Which intermediate is formed during the oxidation process of D-Glucose?

    <p>1-2 Endiol</p> Signup and view all the answers

    Which solution contains a potential carcinogen and requires special care during handling?

    <p>Glucose oxidase with γ-dianisidine hydrochloride</p> Signup and view all the answers

    Which solution is essential in determining the calibration curve for detecting blood glucose?

    <p>Copper Sulphate Solution</p> Signup and view all the answers

    What is the primary purpose of adding Na2WO4 and H2SO4 during the protein precipitation process?

    <p>To precipitate proteins from serum</p> Signup and view all the answers

    What is the first step in the process of protein precipitation from serum?

    <p>Addition of Sodium Tungstate solution</p> Signup and view all the answers

    Which solution is used as a blank in the blood glucose determination?

    <p>CuSO4 - Na2SO4 solution</p> Signup and view all the answers

    During spectrophotometric analysis, what is the wavelength utilized for reading absorbance?

    <p>650nm</p> Signup and view all the answers

    What is the concentration of glucose in Working Standard 2 after dilution?

    <p>200mg/100ml</p> Signup and view all the answers

    Why is it important to cool the test tubes after boiling for 10 minutes?

    <p>To stop the reaction from proceeding</p> Signup and view all the answers

    What role does Harding's solution play in the assay?

    <p>It acts as an anticoagulant.</p> Signup and view all the answers

    What is the first step when preparing the serum for glucose determination?

    <p>Add sodium tungsten to the serum</p> Signup and view all the answers

    What is the final volume to which test solutions are diluted after the addition of distilled water?

    <p>25ml</p> Signup and view all the answers

    Study Notes

    Reagents for Glucose Determination

    • Glucose Oxidase Reagent:
      • Contains 0.5 ml glucose oxidase, 5 ml peroxidase solution, and 1 ml of 0.5% o-dianisidine-HCl in acetate buffer.
      • Stores well in the refrigerator for at least two weeks.
    • Standard Glucose Solution:
      • Prepared by dissolving 100 mg of glucose in 100 ml of 0.1% benzoic acid.
      • This standard solution is equivalent to 100 mg glucose/100 ml.
      • Glucose solutions are stable indefinitely at room temperature.
    • Protein Precipitation & Removal Reagents:
      • Section A: Zinc Sulphate, Sodium Hydroxide, Distilled Water.
      • Section B: Isotonic Sodium Sulphate, Sodium Tungstate, 2/3 N Sulphuric Acid.
    • Glucose Determination Reagents:
      • Harding's Solution: Alkaline Anticoagulant Medium.
      • Nelson's Solution: Contains Arseno-molybdenate (toxic).
      • Copper Sulphate Solution: (CuSO4).
      • Standard Glucose Solutions (WS):
        • Low Standard (WS1): Diluted 5.0ml of stock Standard Solution to 200ml with CuSO4 - NaSO4 solution (equivalent to 100mg/100ml of blood).
        • High Standard (WS2): Diluted 10.0ml of stock Standard solution to 200ml with CuSO4-NaSO4 solution (equivalent to 200mg/100ml of blood).

    Safety Precautions

    • Glucose Oxidase Reagent:
      • Contains γ-dianisidine hydrochloride which is a potential carcinogen. Do not pipette or spill.
    • Standard Glucose Solutions:
      • Contains Benzoic Acid, a skin irritant. Flush exposed areas with water if spilled.
    • Zinc Sulphate:
      • Can cause skin irritation. Flush exposed areas with water if spilled.

    Glucose Determination Method (Section B)

    • Method Overview:
      • Uses a colorimetric method with Nelson's reagent to measure glucose concentration.
    • Steps:
      • Label 4 test tubes: B, S1, S2, and T (Blank, Standards 1 & 2, and Test).
      • Add Harding's solution, CuSO4 solution, supernatant, and appropriate standard solutions as per the table.
      • Place tubes in a boiling water bath for 10 minutes.
      • Allow tubes to cool.
      • Add 2 ml Nelson's arseno-molybdenate solution to each tube.
      • Dilute to 25 ml in a volumetric flask with distilled water.
      • Let the tubes stand for 5 minutes.
      • Read absorbance in a spectrophotometer at a wavelength of 650 nm.

    Glucose Oxidation Reaction

    • Glucose is oxidized to gluconic acid.
    • The oxidation reaction involves the following steps:
      • Glucose is converted to a 1-2 endiol intermediate.
      • Cupric ions (Cu2+) are reduced to cuprous ions (Cu+).
      • Cu2+ (Cupric ions) react with the endiol intermediate to form Cu2O (Cuprous Oxide) and gluconic acid.
      • Sodium sulfate (Na2SO4) is added to prevent reoxidation of Cu2O, ensuring that the Cu2+ ions are used solely in the oxidation reaction.
    • Arseno-molybdenate (Nelson's reagent) reacts with the cuprous oxide (Cu2O) forming a green-blue "molybdenum blue" which is measured spectrophotometrically at 650nm.

    Additional Notes

    • Protein precipitation and removal:
      • Performed using different reagents in Sections A and B.
      • Aims to remove proteins from the serum to obtain a clear supernatant for the glucose determination.
    • Blood Dilution:
      • In Section A, the blood is diluted 25 times during protein precipitation.
      • Remember to multiply the final glucose concentration by 25 to get the actual blood glucose concentration.

    Studying That Suits You

    Use AI to generate personalized quizzes and flashcards to suit your learning preferences.

    Quiz Team

    Related Documents

    Practical 2: Determination of Serum Glucose PDF

    Description

    This quiz explores various reagents used in the determination of glucose levels, including glucose oxidase, standard glucose solutions, and protein precipitation agents. Additionally, it covers the preparation and stabilization of these reagents for effective laboratory use. Test your knowledge on this essential aspect of biochemical analysis.

    More Like This

    Use Quizgecko on...
    Browser
    Open
    Browser
    Browser