Chromatography 1

Question 1

What is chromatography?

Answer 1

process of separation of components from a mixture of compounds

Question 2

How does separation occur?

Answer 2

separation occurs by placing a sample onto a stationary phase
- usually solid but can be liquid

mobile phase is passed through or over the system

sample components move by a process known as elution

Question 3

What is elution?

Answer 3

process of extracting one material from another by washing it through a chromatography column

components having differential rates of migration will elute at different times

Question 4

How is the rate of migration determined?

Answer 4

rate of migration of components through the stationary phase is determined by its distribution ratio, D.

D = Cs/Cm
Cs - solute concentration in stationary phase
Cm - solute concentration in mobile phase

measure of relative affinity in each phase

Question 5

What does a low and high distribution ratio tell you?

Answer 5

high distribution ratio (D) - means it is more soluble in the stationary phase = elutes slowly

low distribution ratio (D) - means it is more soluble in the mobile phase = elutes faster

Question 6

What is sorption and desorption?

Answer 6

sorption
- process where solutes are transferred from the mobile to the stationary phase

desorption
- process where solutes are transferred from the stationary phase to the mobile phase

Question 7

What are the different sorption isotherms in chromatography?

Answer 7

adsorption
partition
ion exchange
size exclusion

Question 8

What is adsorption?

Answer 8

is a surface effect

occurs due to electrostatic interactions between compound and the surface of the stationary phase

Question 9

What is partition?

Answer 9

applied to bonded phases

• when liquid is coated onto a solid to form a station phase
• sorption and desorption occur between the liquid coated onto the stationary phase and the liquid mobile phase

is analogous to solvent extraction

Question 10

What is ion exchange?

Answer 10

solute ions in the mobile phase can exchange with similarly charged ions in the stationary phase

Question 11

What is size exclusion?

Answer 11

no specific interaction between the mobile and stationary phase

stationary phase consists of porous silica or polymer gel containing holes or pores

• larger molecules are unable to get through so pass down faster
• smaller molecules spend time in the holes and pass slower

Question 12

What is a Gaussian Profile?

Answer 12

particles of the same component will not move at the exact same speed
- particles will form a broad band with some particles eluting faster or slower

Question 13

How can the quality of separation be assessed?

Answer 13

efficiency - how sharp the peaks are

resolution - separation of peaks

Question 14

What is efficiency? How can it be calculated?

Answer 14

efficiency (N) is an indication of how sharp the peaks are

N can be calculated using the theoretical plate concept
- the stationary phase is considered a number of small chromatography plates lined up one after the other

Question 15

What is plate height?

Answer 15

plate height - H
height of the theoretical plates making up the stationary phase

H = L/N
L - column length
N - efficiency

Question 16

What is resolution? How can you tell if peaks are well resolved?

Answer 16

a measure of how well separated the peaks are

if R is greater than or equal to 1.5 then the peaks are well resolved

Question 17

How can resolution be improved?

Answer 17

change the solvent - mobile phase

change the column - stationary phase

Question 18

What is retention factor?

Answer 18

measure of retention ability of chromatographic system for a given substance

calculated with respect to an unretained compound
- one that pass through the column with no interaction with the stationary phase

Question 19

What is retention time?

Answer 19

time since the run has started

- time spent in the column