Chromatography Flashcards
What is the Mobile phase?
Solvent that caries the soluble components of the mixture
If a sample moves faster then another.
How does the relationship between the phases differ?
The sample that moves faster has a stronger affinity for the mobile phase.
If a sample moves slower then another.
How does the relationship between the phases differ?
The sample that moves slower has a stronger affinity for the stationary phase.
-usually components are hydrogen bonded to the stationary phase
What is the Stationary phase?
A medium that attracts the components in the mixture, resisting motion against the mobile phase
How are substances separated by chromatography? (3)
- Suitable stationary & mobile phases are choses
- The balance between affinity for the mobile and stationary phases is different for each component in the mixture
- Different components move at different rates, and are separated from one another over time
Why will different substances show different Rf Values?
Explain why the differences occur (3)
- All Molecules are bonded differently & have different polarities
-More polar bonds gives more affinity to the stationary phase and a smaller Rf Value
-Hydrogen bonding and dipoles are attracted more strongly to the stationary phase
What does TLC stand for?
What is the Stationary phase?
What is the Mobile Phase?
Thin Layer Chromatography
Glass, plastic or metal plate coated in silica (SiO2), alumina (Al2O3) or both.
Mobile phase dependant on the possible components of the micxture
What are the advantages of TLC over Paper Chromatography?
Runs Faster.
Smaller amounts of mixture can be separated
Plates are more robust then paper
How do you analyse the results of TLC via UV Light? (4)
-Add fluorescent dye to silica/alumina
-Shine UV light onto plate
-Colourless spots on the chromatogram will block any glow from the fluorescent dye
-Draw round the spots to mark where they are
How do you analyse the results of TLC via Iodine? (2)
- Place chromatogram in a sealed jar with iodine crystals
-Iodine vapour (locating agent) sticks to the colourless spots turning them purple
How do you analyse the results of TLC via Ninhydrin? (2)
What can Ninhydrin analyse?
-Spray chromatogram with heated ninhydrin (developing agent)
-Ninhydrin turns amino acid spots from colourless to purple
-Amino acids
How do you calculate Rf Values
Distance moved by spot / distance travelled by solvent front
Both measured from the baseline
What is an Rf Value?
Retention Factor
A ratio of the rate of movement between the solvent and components within the mixture
How do you confirm the identity of a substance from an Rf value? (2)
-Compare the Rf Value to accepted values of Rf for that substance
- make sure the Rf for the accepted value is the same solvent and set up as your own
What is Column Chromatography? (4)
A glass column pack with silica or alumina which acts as the stationary phase
The mixture and solvent (mobile phase) is run through the column continuously
Different compounds in the mixture run through the column at different rates - producing different retention times
Each pure separated component can be collected separately at different intervals
What is the mobile phase in Column Chromatography?
Solvent that is added to the top and runs down the column
- Called Eluent
What is the stationary phase in Column Chromatography?
Silica Alumina or resin packed into a glass column
Draw a Diagram of Column Chromatography
What are the Advantages of Column Chromatography?
More then one eluent can be used - leads to better separation then TLC
Large amounts of mixture can be separated and collected then TLC
Draw a Diagram for GC
What is the Stationary Phase for GC? (3)
-Silica Or Alumina powder coated with oil
-Packed into a long thin Capillary Tube
-Coiled and placed into a warming oven
What is the Mobile Phase for GC?
Inert Carrier Gas - such as N2, Ar or He
What are the advantages of GC?
Very Sensitive instrument - can detect traces of substances
(can link oil pollution to specific ships, and detect certain types of chemicals in foods)
What are some of the common uses of GC?
Blood/Urine alcohol & drug testing - breathalyser test
How can you use GC or GCMS to identify Substances? (3)
- Separate mixture using GC, and record retention time
- test each component with an MS spectrometer
-Use combination of the Fragmentation pattern/ molecular ion peak graph from the MS with the retention time from GC.
- To compare with accepted values of known substances .
How does GCMS work?
GC is run, separating the mixture and recoding each of the components retention time
Separated components are then run through an MS spectrometer.
Fragmentation pattern/ molecular ion peak confirms the identity of the substance
Which organic substance will have a shorter retention time?
Aldehyde or Alcohol (4)
Aldehyde
-Has a Less of a polar bond then an alcohol (more soluble)
- it adsorbs less strongly to the stationary phase.
- so moves through the column/plate at a faster rate due to less attractive forces with the stationary phase
Plastic gloves prevents contamination of TLC plate
Adding developing agent at a depth below 1cm^3 - if the solvent is too deep it will dissolve the mixture from the plate
Solvent doesn’t need to rise to the top as an Rf value can be calculated regardless
Fume cupboard is used as the solvent is toxic
Amino acids have different bonds and polarities
Therefore have a difference balance between the affinities for the mobile and stationary phases
Resulting in different retention Times
