Class 2 Flashcards
What are the two phases in all types of chromatograph?
- Stationary phase
2. Mobile phase
What is the stationary phase in chromatography?
Substance that supports the mixture allows compounds to be retained. Usually a column of silica gel or beads.
What is the mobile phase in chromatography?
Fluid that carries mixture of compounds to be separated
What is size exclusion chromatography?
Separates compounds based on molecular size.
Used to separate full proteins (large) from peptide fragments (small).
In size exclusion chromatography, which compounds will elute first?
Larger compounds because smaller compounds typically must travel through porous beads in the column whereas large molecules just go around the beads.
What is thin-layer chromatography (TLC)?
Separates compounds based on polarity.
Used to separate small amounts of solids of high BP liquids using a column of silica (polar!).
Describe silica gel.
Silica gel is POLAR and thus can H-bond to some compounds.
Describe the movement of polar versus nonpolar molecules in TLC.
The more polar the compound it is, the more it interacts with the plate and thus does not travel as far.
Nonpolar molecules get easily pushed down the plate by the solvent.
What is the Rf value?
in TLC,
Rf = migration distance (spot) / migration distance (solvent front)
Thus it will ALWAYS be a number
What is a cute trick to remember polar vs nonpolar movement in TLC?
Polar is lower and slower
What is column chromatography? What can it be used for?
Same principle as TLC but in a large column and used to separate LARGE amounts of solids or high BP liquids.
It can be used to monitor a reaction–to see if you still have different compounds or if the reaction has gone to completion, etc.
What is HPLC? How does it work?
High Performance Liquid Chromatography.
Liquid is forced through a column under high pressure so that its is a more efficient version of column chromatography. You can swap out different kinds of columns depending on your compound type.
What is ion exchange chromatography?
Separates compounds based on difference in charge states (+/-, or neutral).
Used to separate mixtures of charged amino acids, proteins, or nucleotides
What is affinity chromatography?
Separates biochemical mixtures based on highly specific lock-and-key mechanisms.
e.g. separate proteins from blood serum or a cell lysate.
What is gas chromatography?
Separates based on differences in volatility/Boiling Point.
Used to separate SMALL amounts of low boiling point compounds.
How do polar vs nonpolar compounds compare in gas chromatography.
Compounds with high boiling points (polar compounds!!) move slower
Compounds with low boiling points (nonpolar!!) will evaporate first.
SO polar/high BP compounds stay in the column longer…Polar is lower and slower still applies
What information can you glean from a gas chromatography output?
- The number of compounds in a mixture is equal to the number of peaks.
- The relative quantity of each compound correlates with area under the peak.
- The volatility/BP is correlated with time. The high BP compounds will peak later on the time scale.
How do you separate a LARGE amount of a compound based on differences in boiling point?
Distillation!
What kind of intermolecular forces keep substances in the liquid (as opposed to gas) phase?
- Hydrogen bonds
- Dipole-dipole forces
- London dispersion forces
Secondary things…
- Heavier molecules
- have higher BPs
More branching = lower BP
What is distillation?
Separation of compounds based on differences in BP.
Used for LARGE amounts of compounds and LOW BP compounds.
In a simple distillation, what is left in the primary glassware? What transfers over to the new glassware and how does this work?
Non-volatile/higher BP compound stays in primary glassware while the volatile, lower BP compound is in secondary glassware.
This relies on a pretty wide difference in boiling point.
When would you do a fractional distillation? What is it useful for?
When BP difference between the two substances is not very large (
What are extractions?
They separate compounds based on their solubility differences in two solvents.
What are the key solubility rules?
- “like dissolves like”:
Polar compounds are soluble in polar solvents (water) and nonpolar compounds are soluble in nonpolar (organic) solvents
- Compounds with
