HPLC Flashcards

0
Q

HPLC is a type of

A

Analytical chrom

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1
Q

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

A

Size exclusion
Reverse phase chrom
Ion exchange chrom

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2
Q

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

A

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

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3
Q

Separates complex mixtures using…

A

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

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4
Q

The columns are packed with

A

Small particles.

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5
Q

The MP is pumped through at a ….

A

Reasonable speed, thus improving efficiancy.

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6
Q

Solvents should be degassed to prevent…

A

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

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7
Q

Can be used for water samples to…

A

Identify presence of small organic contaminants.

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8
Q

HPLC columns have an internal diameter of

A

3 mm to 10 mm

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9
Q

Columns contain spheres of…

A

High purity silica, packed into a dense matrix.

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10
Q

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

A

Better separation efficiancy.

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11
Q

Van D eq: the B value is…

A

Small in HPLC

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12
Q

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

A

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

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13
Q

When the MP is pumped, the sample is…

A

Loaded into the column.

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14
Q

Silica is a good packing material because…

A

It has excellent absorbent properties.

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15
Q

The column packing separates components bun

A

Chemical or physical properties.

16
Q

HPLC is better at separation and analysis of

A

Non volatile compounds.

17
Q

Temperature is critical because (2)

A

High temps may affect stability of analytes.

Retention times are temp dependant.

18
Q

Not all detectors respond to all…

A

Compounds.

19
Q

The signal produced is directly proportional to…

A

The amount of substance present - Quantitive measurement.

20
Q

Difference types of detectors: (5)

A
Photodiode array.
Mass spec.
Fluorescence.
Uv vis.
Universal.
21
Q

How does a uv vis detector work?

A

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.

22
Q

A disadvantage to uv vis detector:

A

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

23
Q

For uv vis, need to know which wavelength…

A

The solute will absorb.

24
Q

Photodiode array: uses polychromatic light passed through eluant….

A

Diff wavelengths passing through sample are absorbed to diff extents.

25
Q

Adv of photodiode array over uv vis:

A

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

26
Q

Dis adv of photodiode array:

A

Low sensitivity. Not all compounds absorb light.

27
Q

A universal detector detects…

A

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

28
Q

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

A

A spectra library and the compound can be identified.

29
Q

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

A

Compared with known retention times for compounds.