Partition Chromatography

Overview of Partition Chromatography

• Partition chromatography separates components of a mixture based on differences in their partition coefficients between an aqueous and immiscible organic liquid.
• The stationary phase may be packed in a chromatography tube or layered on a plate in the form of sheets.
• The mobile phase may be gas, liquid, or vapor, and flows in a counter-current manner to the stationary phase.
• The method is largely replaced by HPLC, but is still used for analysis of lipophilic substances.
• Stationary phases include silica gel, which can be non-polar or polar depending on the type of chromatography being used.
• Paper chromatography is a popular method for fractionation of some groups of substances, using cellulose paper as the stationary phase.
• Efficient development in partition chromatography is achieved by using analytical grade chemicals, deionized water, maintaining constant composition and temperature of the mobile phase, and selecting suitable solvents.
• Equilibration is achieved using the organic-rich phase as the developer and water-rich phase to saturate the atmosphere around the paper with its vapor.
• Resolution of complex mixtures can be improved by adjusting the pH of the developer by addition of specific reagents.
• Spot detection can be accomplished using physical or chemical methods, including enzymatic and biological methods.
• Identification of substances involves determining the retardation factor or Rf value, which is always less than 1.
• Factors affecting Rf values include variations in the stationary phase, mobile phase, temperature, method and direction of development, sample size and concentration, and presence of impurities. Chromatographic artifacts can also occur.