Seperation Techniques Flashcards
What is an Extraction
The transfer of a solute from one phase to another
Used to isolate or concentrate the analyte
The equilibrium determines how efficiently the solute will pass between two phases
What factors Affect Extraction Efficiency?
Polarity - Polar solute favor polar solvents
Type of Compound - Hydrophilic or hydrophobic
pH - ex. As pH increases, the efficiency increases
How Does Chromatography differ from Extractions?
Multiple analytes of a mixture can be separated using chromatography.
Chromatography also uses two phases.
What is the Mobile Phase in Chromatography
The mobile phase is the solvent that moves through the column.
It is a liquid or a gas.
What is the Stationary Phase in Chromatography
The stationary phase is the medium in the colum that the mobile phase passes over.
It is a viscous liquid or a solid.
What factor allows analytes to separate?
Interactions between the analytes and the stationary phase.
Analytes with strong interactions will be slower to pass through the colum that analytes with weaker interactions.
Define Eluent and Eluate as they relate to Elution in Chromatography
The fluent is the fluid entering the column, and the eluate is the fluid leaving the column
What are the two main types of Columns can be used for Chromatography?
Packed Columns are filled with particles of the stationary phase
Open Tubular Columns are capillaries where the stationary phase coats the outside wall.
Describe the difference between Porous Layer Open Tubular (PLOT) Columns and Wall Coated Open Tubular (WCOT) Columns
PLOT is a thin layer of a porus material such as an organic polymer like polystyrene or an inorganic compound like silica
PLOT has a higher sample capacity, and is stable over a larger pH range
WCOT have a lower sample capacity and a smaller diameter. Fewer dilution are generally needed to prepare the sample.
Explain Adsorption Chromatography
The stationary phase is solid
The mobile phase can be liquid or gas
Solute is adsorbed, meaning it interacts with only the surface of the stationary phase
Used in drug testing and urine samples
Explain Partition Chromatography
The stationary phase is a highly viscous liquid bonded to a solid surface
The mobile phase is gas
Solute equilibrates between the gaseous mobile phase and the stationary liquid
Commonly used in gas chromatography
Explain Ion-Exchange Chromatography
The stationary phase is a solid resin with anions or cations covalently attached to it
The mobile phase is a liquid
Solute ions of opposite charge are attracted to the stationary phase, while ions of the same charge pass through. Oppositely charged ions must be eluted at the end.
Used in protein analysis because amino acids are charged
Explain Molecular Exclusion Chromatography
(also known as Gel Filtration or Gel Permeation Chromatography)
Stationary phase is a porus gel
Mobile phase can be a liquid or a gas
Separated solute by size, larger molecules pass through first because smaller molecules get trapped
Solute and stationary phase do not interact
Describe Affinity Chromatography
The most selective type of Chromatography
Specific interactions between the solute and stationary phase occur. For example the stationary phase may be the antibody for a specific protein which immobilize it, allowing it to separate and than be eluted.
What factors are used to determine resolution?
How do they relate to efficiency?
Resolution = delta time/ average width
More time between peaks means there is better separation
Narrow peaks also mean there is better separation
How does Diffusion influence the Efficiency of Chromatography?
Diffusion is the tendency for solute to move from areas of high concentration to areas of low concentration
This increases the range of a solute and decreases efficiency.
Selecting an appropriate stationary phase and mobile phase can minimize this
How does Plate Theory affect Chromatography Efficiency
The greater the number of plates, the more times the analyte interacts with the stationary phase, the greater the efficiency is
What factors affect the Resolution of Chromatography
Number of theoretical plates
You can increase this by increasing the column length
Separation factor - the velocity of the solute traveling through the colum
In Gas Chromatography, you can increase this by changing the stationary phase
In Liquid Chromatography, you can change either the stationary or mobile phase
What Types of Detectors can be used with Chromatography
Mass Spectrometer
Fourier Transform Infrared
Ultraviolet
Simple Chromatography does not require a detector
Explain High Preformance Liquid Chromatography
The Mobile phase is pumped through the column
The sample is added by an autosampler injector
The column mag have a guard colum to protect it from any impurities
Once the sample passes through the column, it reaches a detector, which is analyzed by a computer, and the waste is collected
Any type of detector may be used
Explain Thin Layer Chromatography
Solid phase is a silica coated plate and mobile phase is liquid
Components of a mixture are identified using the Retnetion Factor (Rf)
Rf = distance from start to center of component/ distance traveled by solvent
