Chromatography Overview and Principles
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Questions and Answers

What does Rf stand for in the context of Thin Layer Chromatography?

  • Retention factor (correct)
  • Reference factor
  • Reactivity factor
  • Radiofrequency

    • In paper chromatography, which of the following phases is the stationary phase?

  • Cellulose filter paper (correct)
  • Liquid solvent
  • Gas phase
  • Water

    • If the solvent has travelled 3.5 cm and the sample has travelled 2.8 cm, what is the Rf value of the sample?

  • 0.75
  • 0.80
  • 0.60 (correct)
  • 1.25

    • Which of the following conditions can Thin Layer Chromatography measure amino acid concentration for?

    <p>Phenylketonuria</p> Signup and view all the answers

    What reagent is used in paper chromatography to detect alpha-amino acids?

    <p>Ninhydrin</p> Signup and view all the answers

    What is the primary purpose of using Thin Layer Chromatography?

    <p>For separation or isolation of biochemical metabolites</p> Signup and view all the answers

    At what point should the paper chromatogram be removed from the solvent in the chromatography process?

    <p>After the solvent front has moved 80 to 90% of the way up</p> Signup and view all the answers

    Which of these conditions is characterized by the body not fully breaking down tyrosine and phenylalanine?

    <p>Phenylketonuria</p> Signup and view all the answers

    What is the main purpose of chromatography?

    <p>To separate closely related compounds from a mixture.</p> Signup and view all the answers

    Which component of a chromatographic system holds the sample being analyzed?

    <p>Stationary phase</p> Signup and view all the answers

    In what way do the mobile phase and stationary phase interact in chromatography?

    <p>They allow continuous redistribution of compounds between both phases.</p> Signup and view all the answers

    Which of the following best describes gel exclusion chromatography?

    <p>A method that separates molecules solely by their size.</p> Signup and view all the answers

    What characterizes thin layer chromatography?

    <p>It employs a stationary phase of solid adsorbents on a glass or metal plate.</p> Signup and view all the answers

    What does the term 'elution' refer to in chromatography?

    <p>The process of passing the mobile phase through the column.</p> Signup and view all the answers

    What drives the separation process in thin layer chromatography?

    <p>The relative affinity of compounds toward the stationary and mobile phases.</p> Signup and view all the answers

    Which of the following properties is NOT typically used in chromatography for separation?

    <p>Electrical conductivity</p> Signup and view all the answers

    Study Notes

    Chromatography Overview

    • Chromatography is an analytical technique separating closely related compounds in a mixture.
    • It is used to separate proteins, peptides, amino acids, lipids, carbohydrates, vitamins, and drugs.

    Physical Basis of Separation

    • Compounds are separated based on differences in physical characteristics.
    • Examples include molecular size, shape, charge, volatility, solubility, and absorptivity.

    Essential Components of Chromatographic Systems

    • Stationary Phase: A porous solid matrix, where the sample dissolved in the mobile phase travels through.
    • Mobile Phase: The mixture of substances (to be separated) dissolved in a liquid or gas.

    General Principle of Chromatography

    • All chromatography types involve a stationary and a mobile phase.
    • The mobile phase moves through the stationary phase, carrying the sample components.
    • Separation happens through continuous redistribution between the mobile and stationary phases.

    Definitions (Chromatography)

    • Eluent: The fluid entering the column.
    • Eluate: The fluid exiting the column.
    • Elution: The process of passing the mobile phase through the column.

    Gel Exclusion Chromatography

    • Separates molecules based on size (sieving).
    • Smaller molecules travel longer paths through the gel matrix.

    Ion Exchange Chromatography

    • Separates ions based on their charges.
    • Anions or cations are attracted to oppositely charged groups on the stationary phase.

    Thin Layer Chromatography (TLC)

    • Stationary Phase: A uniform layer of silica gel, cellulose, or alumina coated on a surface (e.g., glass).
    • Separation occurs based on the relative affinity of compounds to the stationary and mobile phases.
    • Compounds with higher affinity for the stationary phase move slower.
    • Rf value: The retention factor, calculated as distance travelled by a spot divided by distance travelled by the solvent.

    Measuring Rf Values

    • Each spot on a TLC plate has a unique Rf value.
    • Rf = Distance travelled by compound / Distance travelled by solvent.

    Paper Chromatography

    • Stationary Phase: Cellulose filter paper.
    • Mobile Phase: Liquid solvent.
    • Separation happens based on the polarity of compounds towards both the mobile and stationary phases.

    Paper Chromatography: Process

    • Cellulose paper is placed in a developing chamber.
    • The sample is spotted on the paper.
    • The solvent moves up the paper, carrying components with it.
    • Compounds with high affinity to the stationary phase move slower.
    • The Rf value can be used to identify compounds.

    Paper Chromatography: Specific Applications

    • Used in identifying amino acids (e.g., using ninhydrin to visualize them).
    • Unknown mixtures can be compared to known standards for identification.

    TLC and Specific Applications

    • Used in Biochemical analysis, such as separating or isolating biochemical metabolites from blood plasma, urine, body fluids, and serum.
    • Used in testing for conditions like phenylketonuria, alkaptonuria, and maple syrup disease.

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    Related Documents

    Chromatography Lecture PDF

    Description

    Explore the fundamental concepts of chromatography, an essential analytical technique used to separate mixtures of compounds. Learn about its physical basis, key components such as stationary and mobile phases, and the general principles that govern the separation process.

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