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Chromatography
Chromatography
55
Biology
Undergraduate 1
03/22/2016

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Term
What is Chromatography?
Definition
The separation of complex materials
Term
How is abundance shown on the chromatograph?
Definition
The area under the peak
Term
How is retention time shown on the Chromatograph?
Definition
On the X axis
Term
How is concentration shown on the chromatograph?
Definition
On the Y axis
Term
What is the Mobile Phase?
Definition
Molecules dissolved in the solvent
Term
What is the stationary phase?
Definition
Molecules adsorbed on the solid packaging
Term
What determines the stationary phase?
Definition
The polarity of molecules
Term
What is a covalent bond?
Definition
A shared pair of electrons that are not shared equally
Term
What is electronegativity
Definition
The ability of an atom to attract the electron density towards itself in a covalent bond
Term
If Charge separation increases, Polarity _______?
Definition
Increases
Term
Give examples of polar molecules
Definition
O-H, C-O, C=O, C-N, N-H
Term
Give examples of non-polar molecules
Definition
C-H C-C
Term
Chromatography only works if the peaks are properly ________?
Definition
Resolved
Term
What is retention time used to identify?
Definition
The molecule
Term
Wht are different affinities caused by?
Definition
Differences in polarity
Term
Is the retention time of the mobile phase long or short?
Definition
Short
Term
Is the retention time of the stationary phase long or short?
Definition
Long
Term
What does HPLC stand for?
Definition
High performance liquid chromatography
Term
What state of matter is the mobile phase in HPLC?
Definition
Liquid
Term
In reversed phase chromatography, what is the stationary phase and why is it used?
Definition
Silica (SiO2) Used because it's easy to make into small particles
Term
What is used instead of silica for the stationary phase in reversed phase chromatography when polar molecules are being analysed?
Definition
Octadecylsilane (ODS)
Chemically modified silica
Term
Why is UV absorption spectroscopy universal?
Definition
Everything will absorb UV light at a short enough wavelength
Term
Name some applications of HPLC
Definition

Identifying drugs

Vitamin levels in food

Sugars in honey and fruit juice

Caffeine levels in drinks

Term
What are the structures in a column chromatographer?
Definition
-Stainless steel tube packed with solid - stationary phase
-Inlet
-Detector
Term
What is retention time?
Definition
The time taken for a component to reach the detector?
Term
What data gives qualitative information and what gives quantitative information?
Definition
Qualitative - time
Quantitative - area under peak
Term
What does time give information on?
Definition
The identity of the substance
Term
What does the area under the peak measure?
Definition
The abundance of the substance
Term
What makes molecules move quickly or slowly?
Definition
Move quickly - if dissolved in solvent
Move slowly - if adsorb onto stationary phase
Term
What determines how much a molecule dissolves and adsorbs?
Definition
The polarity of it
- different affinities for mobile and stationary phase
Term
Why do polar molecules dissolve in the solvent?
Definition
The solvent is polar
Term
Why do non polar molecules adsorb to packing?
Definition
The packing is non polar
Term
What does it mean if the graph is well resolved?
Definition
The peaks are separate with clear gaps
Term
Why are small particles of stationary phase used?
Definition
Increases surface area molecules can adsorb onto
Term
What is the disadvantage of using SiO2 as the stationary phase?
Definition
Very polar so polar molecules stick there for a long time
Term
How is ODS modified to make it less polar?
Definition
Coated with non polar hydrocarbons to balance charge
Term
What structures does a HPLC chromatographer have?
Definition
-Stainless steel tube packed with stationary phase
-Solvent reservoir and pump
-Mobile phase
-Injection valve
-Detector
Term
What is used as a mobile phase in HPLC?
Definition
A solvent more polar than the stationary phase
Term
Which is the most and least polar out of methanol, water and acetonitrile?
Definition
Most - water
methanol
Least - acetonitrile
Term
What is the purpose of a loading loop?
Definition
So the solvent's path isn't disrupted by analyte
Term
What is used as a detector and why?
Definition
UV absorption spectrometry, because it's universal
Term
What is the mobile phase in gas chromatography?
Definition
An inert gas
Term
What is the stationary phase in gas chromatography?
Definition
-Packed column
-Capillary column
Term
What is a packed column in gas chromatography like?
Definition
-Stainless steel
-1m long
-5mm in diameter
-Particles of solid packed inside
Term
What is a capillary column in gas chromatography like?
Definition
-Glass
-3m long
-1/10mm in diameter
-Plastic (polyimide) coating
-Stationary phase layer on inside of tube
Term
Where does gas chromatography take place?
Definition
In an oven, up to 300oC
Term
What does increasing the temperature when gas chromatography is taking place do?
Definition
Extends the range of things analysed
Term
What stationary phases are used for gas chromatography and what for?
Definition
Squalene - non polar, used for hydrocarbons, oils
Diethylene Glycol succinate - moderately polar, used for fatty acids, methyl esters
Polyethylene glycol - polar
Term
How polar should the stationary phase be in relation to the sample?
Definition
Same polarity
Term
How is gas chromatography carried out?
Definition
-Sample injected into vapourisation chamber and moves down with carrier gas
-Substance added with analyte to get internal standard
-Concentration of unknown is determined by comparing peak areas with peak area on internal standard
Term
What is the detector in gas chromatography?
Definition
Flame ionisation detector
Term
How does a flame ionisation detector work?
Definition
-Ions burn in different ways
-detected as electric current
Term
What are the advantages of a flame ionisation detector?
Definition
-Can measure low concentrations
-Will detect anything that burns with a good sensitivity
Term
What are the applications of gas chromatography?
Definition
-GS-MS (gas spec - mass spec)
-Disease diagnosis
Analysing:
-Pesticidal residue
-Molecules with low molecular weight - volatile
-Alcohols
-Hydrocarbons
-Fatty acids from triglyceride lipids
Term
How can gas chromatography be used to diagnose diseases?
Definition
By identifying compounds in breath, body fluids
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