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Chromatography Dr.Tawfeq A. Al-Howiriny Associate Professor

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Presentation on theme: "Chromatography Dr.Tawfeq A. Al-Howiriny Associate Professor"— Presentation transcript:

1 Chromatography Dr.Tawfeq A. Al-Howiriny Associate Professor talhowiriny@yahoo.com

2 Adsorbents Adsorbents ► Silica gel ► Silica gel is the most commonly used adsorbent in TLC studies. It is prepared by Hydrolysis of sodium silicate to polysilicic acid, which on further condensation and polymerization yields silica gel material. The synthesis can be controlled so as to yield silicas of high purity and of the required specification with regard to pore volume pore size and specific surface area. A binder is commonly added to the silica gel to confer greater mechanical strength to layer and to enhance adhesion to the baking plate.

3 Adsorbents Adsorbents ► The suffix G is used universally to denote silica with gypsum binder, namely calcium sulphate hemi – hydrate (CaSo4(0.5H2O)). The presence of calcium ion does not affect most separations, however there are available silica gel which adhere sufficiently without the use of the binder. Another binder which has found a limited use is starch, though it places restriction on the use of corrosive location agents. The binder is present at about 10% w/w. The resolution and separating efficiency achieved are dependent, as in other forms of chromatography, upon particle size and Particle size distribution. The resolution improves as particles size becomes smaller and particle – size distribution narrower. The trend towards smaller and more uniform particles continue and the silica gel now commonly in use for TLC studies has a mean particle size of 15 µ m with particle size ranges of 5-40 µ m.

4 Adsorbents Adsorbents ► Commercially available pre coated plates have a mean particle size of 10 µ m with a correspondingly narrower particle size range, with thinner layers of 250 µ m for increased speed and resolution. The newer technique of high- performance TLC (HPTLC) uses silica gel with particle size of 5-6 µ m ► Plates are available with fluorescence indicator commonly a phosphor, which emits a green fluorescence when irradiated with UV light (Hg lamp) of 254nm. The absorbing substance appears as a dark spot against the green fluorescence background due to quenching of the fluorescence.

5 Adsorbents ► The sorption mechanism predominant on silica gel is adsorption and the plates with suitable choice of eluent can be used to separate the neutral, basic and acidic hydrophilic substances, the silica gel can be farther modified to afford reverse – Phase chromatography. Reverse – phase plates have a long chain hydrocarbon impregnated into the support giving packings of defined composition and uniforms coverage which behave similarly to octadecysilane (ODS) modified silica( Chapter 6). These materials are of the use for the analysis of lipophilic substances, fats and waxes,steroids and fat-soluble vitamins and dyes.

6 Adsorbents ► Silanised Silica Gel ► This material is separated by treating silica gel with silanizing agent, for instance dimethyldichlorosilane. The silanol groups of the silica gel are mainly transformed to dimethyle silyl groups with little adsorptive capacity. ► This silanized material can be used to advantage as a lipophilic stationary phase in reverse – phase chromatography.

7 Adsorbents ► Alumina ► Alumina (aluminum oxide) can be synthesized to the same degree of purity and specification as silica gel by a series of non- uniform dehydration processes on various crystalline modifications of aluminum hydroxide. Further more the reaction conditions can be adjusted to produce aluminum oxide with a surface either acidic, basic or neutral; it can be used with or without a binder and the use in the latter form is more common than with silica.


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