PAG06 = CHROMATOGRAPHY

Question 1

What is chromatography used for?

Answer 1

used to separate a mixture into the different

biological molecules​.

Question 2

What is the stationary phase?

Answer 2

mobile phase passes over it eg = TLC plates (plastic sheet with silica gel) OR Chromatography paper (paper made of cellulose)

Question 3

What is the Mobile phase?

Answer 3

travels up

the stationary phase​ and carries the biological molecules with it. eg = liquid/gas

Question 4

What will happen in chromatography?

Answer 4

Each biological molecule will be adsorbed​ onto the surface of the TLC by
different strengths​. Molecules which are adsorbed more ​will travel up the plate slower​,
meaning that different molecules will travel at different speeds up the plate​, being
separated​ as they do.

Question 5

What is adsorption?

Answer 5

= where molecules bond to the surface of a substance

Question 6

How do you set up chromatography for separating amino acids?

Answer 6

1. Draw a pencil line at the bottom of chromatography paper​.
2. Put a concentrated spot of the mixture of amino acids on the paper.
3. Add a small amount of solvent​ (butan-1-ol, glacial ethanoic acid and water) to the
   beaker. Put the chromatography paper in​ and ensure the pencil line is above the
   solvent line​.
4. Place a watch glass over the beaker to stop the solvent evaporating​. This should be
   done in a fume cupboard.

Question 7

How does chromatography work for separating amino acids?

Answer 7

5. As the solvent moves up the paper, the amino acids separate
6. When the solvent almost reaches the top, take the paper out and mark the solvent
   front ​with a pencil.
7. Leave the paper to dry.
8. Amino acids aren’t coloured so spray the amino acids with ninhydrin solution,
   which makes them go purple​. This should be done in a fume cupboard.

Question 8

How do you calculate RF value?

Answer 8

Measure the distance the solvent moved​ and the distance each spot moved up the
plate​.
10. Calculate the Rf value​ for each spot by dividing the distance travelled by the spot
by the distance travelled by the solvent​ (note: you should always get a value less
than 1!!

Question 9

What is the formula for RF value?

Answer 9

value = distance travelled by pigment (divided by) distance travelled by solvent