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  Section: Practical Skills in Chemistry » Instrumental techniques
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Quantitative analysis

Instrumental techniques
  Basic spectroscopy
    Introduction to spectroscopy
    UV Ivisible spectrophotometry
    Fluorescence spectrophotometry
    Phosphorescence and luminescence
    Atomic spectroscopy
  Atomic spectroscopy
    Atomic Absorption Spectroscopy
    Atomic Emission Spectroscopy
    Inductively coupled plasma
    Decomposition techniques for solid inorganic samples
  Infrared spectroscopy
  Nuclear magnetic resonance spectrometry
    1H-NMR spectra
    13C-NMR spectra
  Mass spectrometry
    Interfacing mass spectrometry
  Chromatography ~ introduction
    The chromatogram
  Gas and liquid chromatography
    Gas chromatography
    Liquid chromatography
    High-performance liquid chromatography
    Interpreting chromatograms
    Optimizing chromatographic separations
    Quantitative analysis
    The supporting medium
    Capillary electrophoresis
    Capillary zone electrophoresis (CZE)
    Micellar electrokinetic chromatography (MEKC)
  Electroanalytical techniques
    Potentiometry and ion-selective electrodes
    Voltammetric methods
    Oxygen electrodes
    Coulometric methods
    Cyclic voltammetry
  Radioactive isotopes and their uses
    Radioactive decay
    Measuring radioactivity
    Chemical applications for radioactive isotopes
    Working practices when using radioactive isotopes
  Thermal analysis

Most detectors and chemical assay systems give a linear response with increasing amounts of the test substance over a given 'working range'. Alternative ways of converting the measured response to an amount of substance are:
  • External standardization: this is applicable where the sample volume is sufficiently precise to give reproducible results (e.g. HPLC). You measure the peak areas (or heights) of known amounts of the substance to give a calibration factor or calibration curve which can be used to calculate the amount of test substance in the sample.
  • Internal standardization: where you add a known amount of a reference substance (not originally present in the sample) to the sample, to give an additional peak in the elution profile. You determine the response of the detector to the test and reference substances by analysing a standard containing known amounts of both substances, to provide a response factor (r), where:

    ⇒ Equation [32.5] r = peak area (or height) of test substance  
    peak area (or height) of reference substance

    Use this response factor to quantify the amount of test substance (Qt) in a sample containing a known amount of the reference substance (Qr), from the relationship:

    ⇒ Equation [32.6] Qt = [peak area (or height) of test substance] × Qr  
    [peak area (or height) of reference substance] r

    Internal standardization should be the method of choice wherever possible, since it is unaffected by small variations in sample volume (e.g. for GC microsyringe injection). The internal standard should be chemically similar to the test substance(s) and must give a peak that is distinct from all other substances in the sample. An additional advantage of an internal standard which is chemically related to the test substance is that it may show up problems due to changes in detector response, incomplete derivatization, etc. A disadvantage is that it may be difficult to fit an internal standard peak into a complex chromatogram.


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