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  • Determination of Blood Glucose

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    Jim Livingston
    704 views14 pages
    This document describes the Folin-Wu method for determining glucose levels in serum. The method involves oxidizing glucose using alkaline copper reagent, which reduces Cu2+ to Cu+. The Cu+ then forms a blue complex with phosphomolybdic acid, which is measured spectrophotometrically. A protein-free filtrate of blood is prepared and reacted with alkaline copper reagent and phosphomolybdic acid. Absorbance readings of the unknown and glucose standards are used to calculate the glucose concentration in mg/mL in the original blood sample. The method provides a chemical means for quantifying glucose that does not require enzymes.

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    Folin-Wu method for determining blood glucose is discussed.

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    This document describes the Folin-Wu method for determining glucose levels in serum. The method involves oxidizing glucose using alkaline copper reagent, which reduces Cu2+ to Cu+. The Cu+ then forms a blue complex with phosphomolybdic acid, which is measured spectrophotometrically. A protein-free filtrate of blood is prepared and reacted with alkaline copper reagent and phosphomolybdic acid. Absorbance readings of the unknown and glucose standards are used to calculate the glucose concentration in mg/mL in the original blood sample. The method provides a chemical means for quantifying glucose that does not require enzymes.

    Copyright:

    © All Rights Reserved
    Flag for inappropriate content
    704 views14 pages

    Determination of Blood Glucose

    Uploaded by

    Jim Livingston
    This document describes the Folin-Wu method for determining glucose levels in serum. The method involves oxidizing glucose using alkaline copper reagent, which reduces Cu2+ to Cu+. The Cu+ then forms a blue complex with phosphomolybdic acid, which is measured spectrophotometrically. A protein-free filtrate of blood is prepared and reacted with alkaline copper reagent and phosphomolybdic acid. Absorbance readings of the unknown and glucose standards are used to calculate the glucose concentration in mg/mL in the original blood sample. The method provides a chemical means for quantifying glucose that does not require enzymes.

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    © All Rights Reserved
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    of 14

    DETERMINATION OF GLUCOSE

    IN SERUM

    D.Jim Livingston,
    Asst.Prof. of Chemistry,
    St. John’s College.
    METHODS
    CHEMICAL METHODS

    ENZYMATIC METHODS
    CHEMICAL METHODS
     Oxidation - Reduction Methods
    * Folin – Wu
    * Somogyi - Nelson

     Condensation Methods
    * orto - Toluidine
    Folin - Wu
    Glucose + Cu2+ Gloconate + Cu+
    pH > 7 , heat

    Cu+ + phosphomolybdate blue


    molybdenium
    complex
    λ = 660 nm
    Follin Wu Method

    OXIDATION
     Glucose is oxidised by Fehling’s solution

     It is a mixture of copper sulphate solution (solution A)


    and alkaline sodium-potassium tartrate solution
    (solution B).
     Cu2+ ions Cu1+ ions
     Cu2O
    Reagents Required:
     Alkaline copper-reagent
    40 gms of anhydrous sodium carbonate (Na2CO3) in 400 mL of
    water and transfer it into a 1-liter flask. Dissolve 7.5 gms of
    tartaric acid and 4.5 gms of CuSO4, which is dissolved in 100 mL
    water and make it up to 1 liter.
     Phosphomolybdic acid reagent:
    5 gms of sodium tungstate + 35 gm of phosphomolybdic acid+20
    mL 10% NaOH+50 mL distilled water. Boil vigorously for 20-30
    min cool and dilute to 350 ml with water. Add 125 ml of ortho
    phosphoric acid and make up the volume up to 500 ml.
     Standard glucose solution:
    100 mg of glucose in 100 ml distilled water (stock) and take 10
    ml from this stock and make it up to 100 ml.
    Preparation of protein free
    filtrate:
     1 ml blood sample, add 8 ml distilled water,
     0.5 ml of 2/3 N sulfuric acid and 0.5 ml of
    10% sodium tungstate solution in a stoppered
    centrifuge tube and mix the contents.
     Centrifuge at 3000 rpm for 10 min and
    collect the supernatant as sample.
     Unknown - 2 ml(1:10) of
    protein free filtrate(blood)
     Standard 2ml glucose
     Blank - 2ml water
     Pipette out 2ml of alk.copper
    and heat for 8 min.

     Allowed it to cool and add


    2ml phosphomolybdic acid.

    U S B  Fill the tubes upto 25ml with


    water.
     Set the wavelength
    420 nm
     Absorbance to zero
    with blank.
     Then read the
    absorbance of
    unknown and
    standard
    Calculations
    Blood glucose (mg/ml) = Absunknown x mg of std. glucose x 100/V. of blood
    Abs Standard

    = Absunknown x 0.2 x 100/0.2


    Abs Standard

    = Absunknown x 100
    Abs Standard
    REFERENCES
     Standard Methods of Clinical Chemistry, Volume 1edited
    by Miriam Reiner Elsevier, 02-Dec-2012
     A Text book of pharamaceutical chemistry, Jayashree
    Gosh, S. Chand Limited 2012.
     https://pdfs.semanticscholar.org/4591/f4ffab9cac51dcace5ffd
    20cfe80b9f75967.pdf
     https://www.biochemden.com/estimation-blood-glucose-level-
    folin-wu-method/

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