Methods of analysis and detection

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 OCR Chemistry A 2815/04 Methods of Analysis and Detection

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OCR A Chemistry

2815/04

Methods of Analysis and Detection

2008 Sherborne School                                  Version 2.0

  • Separation techniques of analysis (PC / TLC / GLC / Electrophoresis)
  • Mass Spectroscopy
  • Atomic emission spectroscopy
  • UV / Visible absorption spectroscopy
  • Combined spectral techniques (NMR / IR / Mass Spectra)

By U6F Jacky Huang 2008/03

A. Introduction to chromatography

All chromatography have the following characteristics:

  1. They all have 2 phases, one is a stationary phase and the other one is mobile phase.
  2. The dissolved compounds – solutes.
  3. There are 2 possible mechanisms, one is partition and the other one is adsorption.

In partition:

The solutes move between the 2 phases. If it is in the mobile phase, the solutes will moves with it. Therefore, if we do spend more time in the mobile phase, it can move further.

In Adsorption:

The stationary phase is usually a polar solid and the solutes are polar molecules. The polar molecules will not enter the stationary phase, but it will hold on the surface of the polar stationary phase.

B. Paper Chromatography - The most basic chromatography

Stationary Phase: Cellulose fiber in the filter paper

Mobile Phase: Liquid Solvent

In paper chromatography, coloured compounds can be separated into many colours, but in some cases, colourless spots are involved. When this happen, chemicals like NINHYDRIN will be sprayed to those substances, and those substances will form coloured complexes with these coloured compounds.

The solutes can be identified in 2 ways:

  1. Comparison to the reference compounds
  2. Calculation of Retardation factor =

D moved by centre of solute spots

                                ____________________________

                                 D moved by front of mobile phase

Sometimes a 2-way chromatography is carried out to ensure that all the solutions are fully separated.

C. Thin layer Chromatography – TLC- Simple, Cheap and Reliable

Stationary Phase: Thin layer of silica

Mobile Phase: Liquid (Silica Gel)

TLC is a very similar technique as paper chromatography, but it is 3 times faster than paper chromatography, works with small amount of sample, a wider range of mixtures separated.

In the stationary phase, the silica is heated to remove water, these acts as a polar solid and the solutes are transported by adsorption in mobile phase.

(When dry, both phases will be likely to occur)

However, silica will attract water and the presence of water becomes the stationary phase and the solutes are separated by partition.

Actual uses: Clinical diagnosis / Forensic testing / Quality control.

D. Gas/Liquid Chromatography - Use for very small sample

Stationary Phase: THE SAMPLE itself

Mobile Phase: Inert Gases or carrier gases 

  • GLC is worked by partition,
  • Flows thru the column of stationary phase
  • Separated due to different solubility,
  • Or volatility,
  • If the stationary phase is non polar  Volatility.
  • Stationary is polar  polar molecules retain Take longer to appear.
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The sample will be detected by the detector and appear on chromatogram.

Chromatogram tells us how much of a component is present, the area under the graph being related to the amount.

So how to Determine the % composition of a mixture by GLC:

The area of a peak = 1/2 (Base × Height)

Therefore, a chromatogram must show all the components and the detector are respond equally to all components.

Green Tea Separation in GLC

【 Retention time: Time between injection and appearance of a component.】

The retention time is specific to particular components for the same ...

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