Chromatography

Question 1

What is Chromatography?

Answer 1

It is a technique used to separate the components of a mixture.

Question 2

What are the two phases of chromatography?

Answer 2

1) Mobile phase -This is where the mixture is dissolved in a liquid and then carried through the stationary phase (Liquid)
2) Stationary phase - This is where the liquid is passed through a structure of different material. (Solid)

You can also have a stationary liquid paired with a mobile gas.

Question 3

How do these the two phases separate mixtures?

Answer 3

The constituents of the mixture will move at different rates through the stationary phase leading to a separation of components. each constituent has a affinity for either the stationary or mobile phase.

Question 4

What type of purposes does chromatography serve?

Answer 4

Analytical or preparation. Or even both.

Question 5

What is the partition coefficient?

Answer 5

It is a relative measure of the concentration of the analyte in the stationary phase compared to the concentration in a mobile phase.

Question 6

What is planar chromatography?

Answer 6

It is when the stationary phase occurs on a plane.

Question 7

what is partition chromatography?

Answer 7

It is testing the solubility of an analyte in your stationary phase against the solubility of the analyte in a mobile phase.

Question 8

What is the resulting pattern in planar chromatography called?

Answer 8

A chromatogram.

Question 9

What are the disadvantages to paper chromatography?

Answer 9

Paper is made from cellulose, which is very coarse and not very absorbent.

Question 10

What is thin layer chromatography (TLC)

Answer 10

It is a planar form of chromatography that is similar to paper chromatography

Question 11

What is the difference between TLC and paper chromatography?

Answer 11

TLC is much more faster, reliable, reproducible and sensitive. TLC uses a glass, aluminium or plastic support in place of paper. The stationary phase used is generally silica, bauxite or cellulose. A binder is used to hold the stationary phase to the support, such as gypsum.

Question 12

What does well-resolved mean in planar chromatography?

Answer 12

It says that is a significant degree of separation from another compound. Increased resolution increases likelihood of being well resolved.

Question 13

What is 2D chromatography?

Answer 13

It is planar chromatography that is rotated 90 degrees after initial separation to get more distinct data. This is useful in more complex mixtures that are not well resolved through one instance of chromatography. A separate solvent may be used in the second run.

Question 14

What is column chromatography?

Answer 14

It is when a glass column (tube) is packed with stationary phase. the stationary phase has both analyte and mobile phase on top and allows gravity to perform the separation. You can also use a pump to separate.

Question 15

What is elution and eluate in column chromatography?

Answer 15

Eluate :It is the portion of solvent and partitioned analyte is removed from the bottom of column.
Elution: is this action.

Question 16

Why is column chromatography so useful?

Answer 16

Because it provides a somewhat pure sample of eluate to be used in spectrometry or spectroscopy.

Question 17

What is absorption chromatography?

Answer 17

Absorption chromatography refers to the stationary phase used, The stationary phase is an absorbent molecule that tends to has a hydroxyl group (strong acid) such as silica or aluminium oxide. This acts as a polar absorber, so analyte constituents that have polar functional groups will be retained more strongly whereas those that dont will be eluated faster.

Question 18

What is partition chromatography?

Answer 18

It is liquid-liquid chromatography, this relies on two non-mixable liquids to separate the analyte by solubilities and therefor their partition coefficient.

Question 19

What is normal and reverse phase partition chromatography?

Answer 19

water(polar) is absorbed into a silica gel to create a stationary phase. and a less-polar liquid is used as mobile phase. Reverse phase swaps the polarity of stationary and mobile.

Question 20

What is the preferred substances for each phase partition?

Answer 20

Polar substances such as amino acids, carbs and water soluble pigments are best resolved by normal phase, lipophilic substances such as fats and fat soluble molecules are best separated with reverse phase.

Question 21

What is a disadvantage to partition chromatography?

Answer 21

The liquid can be stripped of the support, but this could be chemically bonded better as a precaution

Question 22

What is hydrophobic interaction chromatography?

Answer 22

This involves the binding of proteins onto the stationary phase to be eluted out. This is achieved by lightly substituting the stationary phase with hydrophobic groups, which is then neutralised by the high concentration salt mobile phase causing the analyte proteins to attach to the stationary phase. The elution of the proteins relies on the lowering of salt concentration causing the proteins to be eluted out.

Question 23

What is ion-exchange absorption chromatography.

Answer 23

This can be done using a positive (cation) stationary phase or a negative stationary (phase). This causes functional groups to be eluted by ionic charge. if you have a positive stationary you can elute positively charge functional groups, and then to elute the more strongly bound analytes we can change the pH

Question 24

What is affinity chromatography?

Answer 24

It involves immobilised biological ligands that selectively bind to a specific analyte or a specific class of analytes depending on the ligand. to elute the analytes out you can change the conditions that affect biological ligands such as heat, ph, temp. To get a specific analyte you use a competitor ligand to compete for bonding.

Question 25

What is size exclusion chromatography?

Answer 25

This uses a stationary phase without reactions, it relies on the small pores similar to the size of the molecules you are testing to filter out the small molecules. in this chromatography the larger molecules are eluted first and often the small molecules get removed.

Question 26

How do you develop of separation?

Answer 26

To decide what type of separation you need you have to consider the mixture, complexity, solubility, polarity and ionic character of the analyte.