Size-exclusion chromatography

**Size-Exclusion Chromatography (SEC)** separates components of mixtures based on size. A mixture can be made of many substances each with different physical and chemical properties. These can be used to differentiate them. SEC allows components that are larger in size to leave the column first while components that are smaller in size will leave the column last.  

A typical column used in SEC has two substances called phases. They interact with the mixture and help force the separation of the mixture. The mobile phase is a solvent which helps carry the mixture down the column. The stationary phase  does not move. Unlike other chromatography methods, the stationary phase used in SEC does not exploit the polarity of each component.  There is no chemical interaction between the stationary phase and a component.  A column could be packed with two types of stationary phase material.  First the stationary phase could be a porous solid. A common porous solid is silica which has a diameter of 5-10µm and  pore sizes ranging from 100-1000Å. Silica provides excellent rigidity and will not move throughout the column. Porous solids are stable at high pressures and temperatures. This allows for the use of a vast variety of solvents. Unfortunately silica particles have a tendency to allow absorb soluble components. They can break down components into atoms. Retention time, how long the component spends in the column, is plotted against molecular weight. The peaks are similar to peaks in liquid or gas chromatography readout. Normally peaks found in liquid or gas chromatography indicates how many components are in a mixture if the number of components is unknown and the relative amount of each component in the mixture within a given range. In the case of SEC these peaks indicate the relative amount of each component (based on molecular weight) within the permeation region.   

## Advantages/Disadvantages
SEC has several advantages and disadvantages. First in SEC the time it takes the mixture to exit the column can be controlled. Depending on the chemical characteristics of the mixture and its components the process could be quick or slow. Second the peaks are well-defined, sharp, and narrow. All characteristics indicate SCE provides good sensitivity. Next there is low mass loss since the sample theoretically is not supposed to come interact with the stationary phase. Last the stability of the stationary phase allows close packing to provide greater separation of components in the sample.  

A clear disadvantage to SEC is with short retention times the peaks overlap. Little information can be extracted from such data. Resolution (how well two peaks can two peaks be distinguished) must be sacrificed if the process is to be shorter in time. SEC does not provide data to distinguish between components of similar molecular mass. For example if component A has molecular mass og  900 g/mol and component B has molecular mass of 950 g/mol, SEC will show one peak in this region and both components will be represented by the same peak.  To have statistically different peaks there should be a 10% difference between molecular weight. Also lumped into this category are enantiomers and diastereomers which are molecules of the same molecular weight with slightly different structures. Last SEC fails to take into account for a component’s viscosity or resistance to flow.

## References
*Principles of Instrumental Analysis, pp 847.
*Principles of Instrumental Analysis, pp 847-8.
*J.  Klein.  “Thalidomide – Contergan.” Web.  2008.  
*P.  Kline; D.  McInnes.  “Summary of Stereoisomerism.”  College of Sana Monica.  Web.  2000.