Unit 5 Flashcards
Elution chromatography
Column is filled with stationary phase .
Sample is added at the inlet and moved over the stationary phase by a mobile phase (ie solvent).
Analyte partitions between the mobile and stationary phase, moving down the column because of the time spent in the mobile phase.
KA = [A]stat/[A]mob
Describe solid-phase micro-extraction
Support coated with a hydrophobic extraction phase (ex.polymer)
Amount of analyte extracted to its concentration in a sample.
Simple, suitable for on-site sampling.
Can pre-concentrate analyte
Larger partition coefficient means
More partitions into the stationary phase
Define components of the partition coefficient
KA = [A]stationary/[A]mobile
Give examples and properties of solid adsorption media
Silica (SiO2): Polar, slightly acidic
Alumina (Al2O3): Polar, neutral or slightly basic
Steps of thin-layer chromatography
Spot a mixture onto plate of silica/alumina (stationary phase)
The bottom big the plate is immersed in solvent, which then travels up the plate by capillary action (mobile phase)
The mixture separated into spots or bands
Visualizing colourless compounds (uv-shadowing, charring, chemical staining
Elution Adsorption chromatography (preparatory technique for chemical synthesis): steps
Column packed with silica gel or alumina
Load sample mixture as narrow band at top of column.
Elute with organic solvent as mobile phase bands for each component separate out
Collect volume fractions
What can separation science do?
Separate and isolate components of the mixture. The amount of each analyte can then be determined.
What can chromatographic techniques mitigate?
Poor intrinsic selectivity of other analytical techniques.
Benefits/downsides of tlc
Quick, simple, low cost, but only qualitative
Block diagram: HPLC instrumentation
HPLC pump (solvents;mobile phase in), injector (sample in), HPLC Column, Detector (waste out), electronics/computer
HPLC pump function
Forces mobile phase through column at high pressure
Injector
Introduces a reproducible volume of sample onto the column
Column
Effects the separation of analytes
Detector function (HPLC)
Produces a measurable signal when analytes elute from the column
Function: inlet and inline solvent filter
Removes particulate matter and air bubbles (ie. Degas) in mobile phase
Pre-column filter
Remove particulate matter introduced with sample
Guard column
Protects analytical column. Removes particulate matter and chemical components that would irreversibly bind to the analytical column
What would happen without HPLC filters?
Top segment of stationary phase in the analytical column would be rapidly deteriorated
Back pressure regulator
Prevents bubbles from forming in the detector cell as the mobile phase exits the column
Draw out an HPLC diagram
See slide 5 under high performance liquid chromatography
Benefits and trade-offs of decreased particle size
Column more densely packed, separation efficiency increase
Tradeoff: very high back pressures. Gravity insufficient to move mobile phase through column. Pumps required
Slide 6 under high performance liquid chromatography
Analytical column
Used for trace analysis and detection
Diameters between 1-10mm and lengths on order of 10-10^2 um
Preparative and semi-preparative columns
Used for purification of synthesized compounds (laboratory scale)
Column diameters and particle sizes are typically about an order of magnitude larger than analytical columns