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    To Separate The Pigments Extract From Spinach Leaves by Paper Chromatography and Determine Their RF Values

    Uploaded by

    Bipul Kumar
    The document describes a procedure to separate pigments in spinach leaves using paper chromatography and determine their Rf values. Key steps include applying a spinach leaf extract to a paper strip, suspending the strip in a solvent mixture in a sealed jar, observing the separation of pigments as colored spots on the paper, measuring the distance traveled by each spot and the solvent front, and using the measurements to calculate the Rf value of each pigment. The results show that carotene had the highest Rf value of 0.98, while chlorophyll b had the lowest value of 0.10.

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    To Separate The Pigments Extract From Spinach Leaves by Paper Chromatography and Determine Their RF Values

    Uploaded by

    Bipul Kumar
    4K views2 pages
    The document describes a procedure to separate pigments in spinach leaves using paper chromatography and determine their Rf values. Key steps include applying a spinach leaf extract to a paper strip, suspending the strip in a solvent mixture in a sealed jar, observing the separation of pigments as colored spots on the paper, measuring the distance traveled by each spot and the solvent front, and using the measurements to calculate the Rf value of each pigment. The results show that carotene had the highest Rf value of 0.98, while chlorophyll b had the lowest value of 0.10.

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    The document describes a procedure to separate pigments in spinach leaves using paper chromatography and determine their Rf values. Key steps include applying a spinach leaf extract to a paper strip, suspending the strip in a solvent mixture in a sealed jar, observing the separation of pigments as colored spots on the paper, measuring the distance traveled by each spot and the solvent front, and using the measurements to calculate the Rf value of each pigment. The results show that carotene had the highest Rf value of 0.98, while chlorophyll b had the lowest value of 0.10.

    Copyright:

    © All Rights Reserved
    Download as PDF, TXT or read online from Scribd
    Download as pdf or txt
    4K views2 pages

    To Separate The Pigments Extract From Spinach Leaves by Paper Chromatography and Determine Their RF Values

    Uploaded by

    Bipul Kumar
    The document describes a procedure to separate pigments in spinach leaves using paper chromatography and determine their Rf values. Key steps include applying a spinach leaf extract to a paper strip, suspending the strip in a solvent mixture in a sealed jar, observing the separation of pigments as colored spots on the paper, measuring the distance traveled by each spot and the solvent front, and using the measurements to calculate the Rf value of each pigment. The results show that carotene had the highest Rf value of 0.98, while chlorophyll b had the lowest value of 0.10.

    Copyright:

    © All Rights Reserved
    Download as PDF, TXT or read online from Scribd
    Download as pdf or txt
    of 2

    Aim:

    To separate the pigments extract from spinach leaves by paper chromatography and determine their Rf values.

    Theory:
     Paper chromatography is essentially a partition chromatography. In paper chromatography the stationary phase is paper. Paper contains
    22% of water molecules absorbed on about 78% of cellulose.
     The separation of the components of the mixture takes place by partitioning of the components between the stationary phase and mobile
    phase. The mobile phase travels through the paper by capillary action.
     The distribution occurs in a definite ratio which represents the characteristic distribution coefficient of the solution.
     The Rf coefficient ration is given by,

    Materials Required:

    1. Whatman’s filter paper


    2. Extract of leaves and extract of flowers
    3. Chloroform/acetone
    4. Methanol/Acetone
    5. Rubber cork fixed with hook in the centre
    6. Glass jar
    7. Rubber cork fixed with hook in the centre
    8. Test tubes
    9. Distilled water
    10. Petroleum ether

    Procedure:

     Take a Whatman filter paper strip and using a pencil draw a horizontal line 4cm from one end of the paper. Then draw another line
    lengthwise (verticallly) from the centre of the paper. Name the point at which the two lines intersect as P.
     Using a fine capillary tube, put a drop of the extract of spinach leaves at the point P. Let it dry in air.
     Put another drop on the same spot and dry again, so that the spot is rich in the leaf extract.
     Pour equal amounts of isopropyl alcohol and distilled water into a chromatographic chamber and mix it well using a glass rod. This is
    used as the solvent.
     Suspend the filter paper vertically in the chromatographic chamber containing the solvent in such a way that the pencil line remains about
    2cm above the solvent level.
     Close the jar with its lid and keep it undisturbed.
     Notice the rising solvent along with the coloured components of the leaf extract.
     After the solvent has risen to about 15 cm you will notice two different spots of coloured components on the filter paper.
     Take the filter paper out of the jar and using a pencil mark the distance that the solvent has risen on the paper. This is called the solvent
    front.
     Dry the filter paper and put pencil marks at the centre of each spot.
     Measure the distance of each spot from the original line and the distance of the solvent front from the original line.
     Calculated the Rf values of different components of leaf extract by using the formula,

    Observation & Result


    .

    Distance travelled by the Distance travelled by the


    Rf value
    SI.No Component component from the original line solvent from the original line
    (P/X)
    (P) (cm) (X) (cm)
    Orange
    1. 11.9 12.2 0.98
    (Carotene)
    Yellow
    2. 3.1 12.2 0.25
    (Chlorophyll a)
    Light green
    3. 1.2 12.2 0.10
    (Chlorophyll b)
    Dark green
    4. 7.2 12.2 0.59
    (Xanthophyll)
    Inference

     Rf value of orange (Carotene) = 0.98


     Rf value of Yellow (Chlorophyll a) = 0.25
     Rf value of Light green (Chlorophyll b) =0.10
     Rf value of Dark green (Xanthophyll) =0.59

    Precautions

     Use good quality pencil for drawing the reference line so that the mark does not dissolve in the solvent in which the chromatography is
    carried out.
     Always use a fine capillary tube.
     Keep the jar undisturbed and covered during the experiment.
     The spot should be small and rich in mixture.
     Allow the spot to dry before putting the strip in the jar.
     Keep the strip erect. Do not let it curl.
     Do not allow the spot to dip in the solvent.

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