6.3.1 Chromatography And Analysis Flashcards
what is chromatography
analytical technique that separates components in a mixture between a mobile phase and a stationary phase
what does separation by column chromatography depend on
balance between solubility in the moving phase and retention in the stationary phase
how does a solid stationary phase separate
by adsorption
how does a liquid stationary phase separate
by relative solubility
what must the mobile phase be
a liquid or a gas
what must the stationary phase be
a solid (as in thin layer chromatography, TLC ) or either a liquid or a solid on a solid support (as in gas chromatography, GC)
how to calculate the rf value in TLC
distance moved by the amino acid/ distance moved by the solvent
what can TLC use
a mixture of amino acids
chromatography method
draw pencil line 1.5cm from the bottom of the paper
capillary tube put a small drop of amino acid on pencil line
roll up the paper and stand it in the beaker
solvent must be below the pencil line
allw to stand for 20 minutes and mark the final solvent level
spray the paper with nihydrin and put in the oven
why won’t some substances separate
as they have very similar rf values
so some spots may contain more than one compound
thin layer chromatography method
wearing gloves draw a pencil line 1cm aboce the bottom of a TLC plate and mark equally placed spots along the line
use a capillary tube and add a tiny drop of each solution to a different spot and allow the plate to air dry
add solvent to a chamber or large beaker with a lid so that is no more than 1cm in depth
place the TLC plate into the chamber or large beaker with a lid so that it is no more than 1cm in depth
place TLC plate into the chamber, level of solvent is below the pencil line, replace lid to get tight seal
when solvent reaches 1cm from top of the plate remove and mark solvent level with a pencil, allow the plate to dry in a fume cupboard
place the plate under UV light to see the spots
draw around the spots lightly in pencil
calculate the rf values
why do you use pencil in chromatography
as it won’t dissolve in the solvent
why do you wear plastic gloves in chromatography
to prevent contamination from your hands to the plate
why do you use a tiny drop in chromatography
too big a drop will cause different spots to merge
why does the solvent depth matter in chromatography
if the solvent is too deep it will dissolve the sample spots from the plate
