HPLC

Question 0

HPLC is a type of

Answer 0

Analytical chrom

Question 1

Can use chromatography based on chemical and physical properties, including….

Answer 1

Size exclusion
Reverse phase chrom
Ion exchange chrom

Question 2

Can be used to analyse classes of compounds, including… (3)

Answer 2

Salts.
Ions and proteins.
Thermally unstable compounds, eg enzymes.

Question 3

Separates complex mixtures using…

Answer 3

Columns that can withstand high pressure and are resistant to a range of chemicals. Eg steel columns.

Question 4

The columns are packed with

Answer 4

Small particles.

Question 5

The MP is pumped through at a ….

Answer 5

Reasonable speed, thus improving efficiancy.

Question 6

Solvents should be degassed to prevent…

Answer 6

Bubbles from forming. These can interfere with valve operation and also cause peaks on the detector.

Question 7

Can be used for water samples to…

Answer 7

Identify presence of small organic contaminants.

Question 8

HPLC columns have an internal diameter of

Answer 8

3 mm to 10 mm

Question 9

Columns contain spheres of…

Answer 9

High purity silica, packed into a dense matrix.

Question 10

Van D eq: a reduced plate height, values of A and C, leads to a…

Answer 10

Better separation efficiancy.

Question 11

Van D eq: the B value is…

Answer 11

Small in HPLC

Question 12

Sample loops adds a known volume of sample into the column through an….

Answer 12

Injector port, this is isolated from the pump and column, until moved to be connected to the column.

Question 13

When the MP is pumped, the sample is…

Answer 13

Loaded into the column.

Question 14

Silica is a good packing material because…

Answer 14

It has excellent absorbent properties.

Question 15

The column packing separates components bun

Answer 15

Chemical or physical properties.

Question 16

HPLC is better at separation and analysis of

Answer 16

Non volatile compounds.

Question 17

Temperature is critical because (2)

Answer 17

High temps may affect stability of analytes.

Retention times are temp dependant.

Question 18

Not all detectors respond to all…

Answer 18

Compounds.

Question 19

The signal produced is directly proportional to…

Answer 19

The amount of substance present - Quantitive measurement.

Question 20

Difference types of detectors: (5)

Answer 20

Photodiode array.
Mass spec.
Fluorescence.
Uv vis.
Universal.

Question 21

How does a uv vis detector work?

Answer 21

Light of particular wavelength is shined through MP flow, containing eluted compounds. The amount of light absorbed is detected. More light absorbed = more solute present.

Question 22

A disadvantage to uv vis detector:

Answer 22

The compound must contain something that will absorb light at the wavelengths used.

Question 23

For uv vis, need to know which wavelength…

Answer 23

The solute will absorb.

Question 24

Photodiode array: uses polychromatic light passed through eluant….

Answer 24

Diff wavelengths passing through sample are absorbed to diff extents.

Question 25

Adv of photodiode array over uv vis:

Answer 25

Wavelengths are recorded simultaneously. Can analyse species that absorb at diff wavelengths.m

Question 26

Dis adv of photodiode array:

Answer 26

Low sensitivity. Not all compounds absorb light.

Question 27

A universal detector detects…

Answer 27

Changes in refractive index. It is less sensitive than uv vis and fluorescence detectors.

Question 28

Mass spec: produces a mass spectrum for every eluted compound, this is compared with…

Answer 28

A spectra library and the compound can be identified.

Question 29

If HPLC doesn’t have a MS, retention time of analytes can be….

Answer 29

Compared with known retention times for compounds.