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Forensic Chemistry > Lecture 4: Basic chromatography > Flashcards

Lecture 4: Basic chromatography Flashcards

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1
Q

What is TLC governed by?

A

Governed by polarity: Will only work if the sample polarity is the same as the stationary phase polarity and different from the mobile phase polarity

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2
Q

What is the stationary phase used in TLC?

A

A solid microparticulate stationary phase bound to a backing (TLC plate)

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3
Q

What is the mobile phase used in TLC?

A

A blend of analytical grade solvents

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4
Q

What is the use of TLC in forensics?

A

-Separation of inks
-Drug screening

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5
Q

What is the method for TLC?

A
  1. Pour 0.5cm of the mobile phase into TLC tank and place lid to equilibriate for 10mins
  2. Draw pencil lid 1cm above bottom of plate (Base line)
  3. Add crosses at equal distance apart
  4. Draw up sample using capillary tube & spot on the crosses
  5. Place the TLC plate in the tank to develop
  6. Mark the solvent front with a pencil line
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6
Q

What does a high affinity for the stationary phase mean?

A

Won’t travel as far up the plate, will be retained for longer

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7
Q

What does a low affinity for the stationary phase mean?

A

Will travel quickly and higher up the plate

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8
Q

How are the Rf values calculated on a TLC plate?

A

Distance travelled by component / Distance moved by solvent front

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9
Q

What do Rf values provide?

A

Tentative identification of an unknown substance

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10
Q

What is the TLC stationary phase backing made from?

A

Silica plat
Can be glass or aluminum

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11
Q

What size are the stationary phase particles on the TLC plate?

A

range from 10-60nm
*Smaller particles = better seperation

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12
Q

How can the solvent strength of the mobile phase be adjusted?

A

A blend of solvents are used

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13
Q

What are locating reagents in TLC?

A

Bind to functional groups or structures allowing them to be coloured

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14
Q

What are examples of locating reagents in TLC?

A

-Ninhydrin: binds to amino acids
-Anisaldehyde: detects steroids
-Vanillin: detects alcohol

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15
Q

What is gas chromatography used for?

A

To separate mixtures of volatile components

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16
Q

How does the sample move in gas chromatography ?

A

The sample is injected via the injector, released into the carrier gas and transported to the column for separation to occur

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17
Q

What is carrier gas in GC?

A

Can be:
-Nitrogen
-Hydrogen
-Helium
*Must be pressurised, purified and inert (Non-toxic, non-flammable)

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18
Q

What gas is used for capillary columns in GC?

A

Nitrogen and Helium

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19
Q

What gas is used for packed columns in GC?

A

Hydrogen

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20
Q

What is the stationary phase used in GC?

A

High boiling point liquids, waxes and oils

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21
Q

What is specific about the stationary phase in GC?

A

specific polarities to analyse different drugs

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22
Q

What is the auto sampler on the GC instrument?

A

Has a syringe and needle
-needle penetrates the GC vial
-sample is drawn up into the syringe and injects into injector

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23
Q

What is the injector on the GC instrument?

A

Releases the sample into the sample inlet, then released into the carrier gas stream and taken to the column

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24
Q

What is the column on the GC instrument?

A

Sits in an oven, where separation occurs

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25
What is a splitless injector?
-All sample is condensed at top of column -Area above column is just above the boiling point of the carrier gas -As temp increased the sample is released into carrier gas stream
26
What's an advantage of the splitless injector?
All of the sample reaches the column for separation
27
What's an disadvantage of the splitless injector?
Can overload the capillary column
28
What is a split injector?
Only 2% of the sample reaches the column, the rest is vented off
29
What's an advantage of the split injector?
Doesn't overload the column
30
What's a disadvantage of the split injector?
Less sensitive due to less sample
31
What's an on-column injector?
-All sample is condensed in a cool zone on top of column -Sample is volatised by programmed heating and released into carrier stream
32
What's an advantage of an on-column injector?
Minimises degradation to components easily degraded by heat
33
What is a flash vaporisation injector?
-Sample is injected into a hot zone, 20-50 degrees above column temp -Sample is then broken down and volatised
34
What is a disadvantage of a flash vaporisation injector?
Can't be used for thermally liable components
35
Which injectors are used for capillary GC columns?
-Split injector -Splitless injector -On-column injector
36
Which injectors are used for packed GC columns?
-On-column injector -Flash vaporisation injector
37
What is the capillary column used in GC?
-Long, narrow tubing made from quartz -Must be wrapped around a housing as can be up to 100m -Small internal diameter, split injector reduces overload
38
Where is the stationary phase on a capillary GC column?
Coats the inner wall, thin layer
39
What is a packed GC column?
-Shorter columns, max 2m -Larger internal diameter -Made of glass
40
Where is the stationary phase in a packed GC column?
packed with a granular solid made from silica particles
41
Which is the more efficient GC column for separation?
Capillary columns due to thinner diameter and longer tube
42
Where are the GC columns stored?
In an electric fan oven
43
What is isothermal programming?
The temperature of the GC oven remains constant throughout analysis
44
What is temperature programming?
The temperature of the oven has programmed changes throughout analysis
45
What is the output of a GC detector?
A chromatogram of the detected separate components
46
What is the retention time?
Time taken to elute from the school
47
What are the universal GC detectors?
-Thermal conductivity detector -Flame ionisation detector
48
What is the thermal conductivity detector?
Detects changes in the thermal conductivity of the carrier gas and compares it to a reference flow of carrier gas
49
What is a flame ionisation detector?
-Carrier gas elutes from column and mixes with hydrogen and is burnt -Produces ions that are collected at a negative electrode -Produces a current that's directly proportionate to the conc of the sample
50
What are the specific GC detectors?
-Nitrogen phosphorus detector -Electron capture detector -Mass spectrometer
51
What is the nitrogen phosphorus detector?
-Detects nitrogen and phosphorus containing groups -A modified FID -Nitrogen detection is increased by 50 -Phosphorus detection is increased by 500
52
What is the electron capture detector?
-Specific for electronegative elements -e.g. nitrogen, sulfur, chlorine, oxygen
53
What is the mass spectrometer detector?
-Most specific -Can positively identify components -Samples break down in GC elute and break down into ions of specific mass to charge ratio
54
What is High performance liquid chromatography?
Separates a mixture into it's components at room temperature
55
How is the sample transported in HPLC?
By a pressurised flow of liquid mobile phase
56
What is normal phase HPLC?
-The mobile phase is non-polar -Stationary phase is polar -Non-polar components have a reduced affinity for stationary phase and elute quicker
57
What is reverse phases HPLC?
-Mobile phase is polar -Stationary phase is non-polar *Most common SP is C18
58
What must be pumped into the mobile phase in HPLC?
Hydrogen gas must be pumped to reduce air and prevent air lock
59
What is the mobile phase used in HPLC?
A mix of high purity HPLC grade solvents
60
What pumps the mobile phase around the HPLC instrument?
Reciprocal pumps ensure the mobile phase is pumped at a constant flow and pressure
61
What is important for separation to occur during HPLC in term of polarity?
The polarity of the sample must be the same as the stationary phase and the polarity of the mobile phase must be different
62
What is solvent programming?
The composition of the mobile phase can be changed over the course of the analysis
63
What are examples of polar HPLC mobile phases?
-Water -Acetic acid -Ethanol -Methanol -Acetone
64
What are examples of non-polar HPLC mobile phases?
-Pentane -Hexane -Cyclohexane -Tolvene -Ether
65
What is the HPLC injector?
-Valve, with or without a sample loop
66
How does the HPLC injector work?
Sample is injected and flows into the sample loop Mobile phase enter and moves the sample through the loop
67
What are the HPLC columns?
-Conventional column -Microbore column
68
What is the conventional HPLC column?
Stainless steel Operating pressure is low: 500-3000 Flow rate is high: 1000-3000ul
69
What is the microbore HPLC column?
Stainless steel Operating pressure is high: 1000-5000 Ideal for HPLC as more sensitive Flow rate is low: 10-100ul
70
What are the HPLC detectors?
-Mass spectrometry -UV-visible absorbance -Fluorescent detector Refractive index detector(Non-specific)

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