chromatography. separation technique that relies on the affinity of components of a mixture to:- –...
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Chromatography
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Chromatography
• Separation technique that relies on the affinity of components of a mixture to:-– A Mobile Phase– A Stationary Phase
• All types of chromatography rely on this– Paper chromatography– thin layer chromatography, – high performance liquid chromatography, – gas chromatography
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Chromatography
• Mobile Phase– This is the phase that is a carrier for the
components as they travel through the stationary phase
• Stationary Phase– This is the phase that is stationary!!
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Types of Chromatography
• Paper Chromatography• Thin layer chromatography (TLC)• High Performance Liquid Chromatography
(HPLC)• Gas Chromatography (GC)
there are more......................
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Paper Chromatography
• Basic technique• Stationary Phase – Paper• Mobile Phase – Usually water• Sample preparation• Good for separating water soluble inks• Limited types of mobile phases can be used
therefore limit to what can be separated• Can be slower • Detection – naked eye or UV light
Paper chromatography.rv
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Thin Layer Chromatography
• Improvement on paper technique• Stationary Phase – silica or alumina based
– Acidic, basic, neutral• Mobile Phase – solvents (varied polarities)• Good for:-
– Monitoring progress of a reaction– Identification of components in a mixture– Purity determination– Conditions for column separation techniques
• Semi-quantative • Can be quicker than paper – phases• Detection – UV light, chemical spray, iodine tank
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Rf Values (retention factor)
• Distance travelled by component
• Distance travelled by mobile phase
• Specific to each plate• Comparable• Each component on
plate has unique Rf value
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High Performance Liquid Chromatography
• Mobile phase – solvents/water/mixes• Stationary phases – silica based/many types for certain
applications• Sample preparation• Precise injection volumes• Composition/flow of mobile phase controlled• Oven temperature• Widely used in analytical laboratories• Quantify components – non volatile mixtures• Retention times
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Gas Chromatography
• Mobile phase – unreactive gas (helium, nitrogen)• Stationary phases – polymer or liquid on an inert
solid support• Sample preparation• Precise injection volumes• Temperature/gas flow control• Widely used in analytical laboratories• Quantify components – volatile mixtures• Retention times
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Columns
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Injector
Autosampler
Detector
Data collection software
Column oven
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AutosamplerData collection software
Column oven Keypad
Injector
Detector
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Applications in Pharma Industry
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Chromatogram
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The Future• Predictive software• Screening• Saving resources