CC1-FINALS-READING CHROMATOGRAPHY AND MASS SPECTROMETRY Flashcards
has long been considered the gold standard for most testing due to its high sensitivity and specificity
Liquid or gas chromatography (GC) coupled to mass spectrometry (MS)
Refers to the group of techniques used to separate complex mixtures on the basis of different physical interactions between the individual compounds and the stationary phase of the system.
CHROMATOGRAPHY
Basic Components of CHROMATOGRAPHY
o Mobile phase
o Stationary phase
o Columns
o Separated components (eluate)
types of MODES OF SEPARATION
- Adsorption
- Partition
- Steric Exclusion
- Ion-Exchange Chromatography
Also known as liquid–solid chromatography, is based on the competition between the sample and the mobile phase for adsorptive sites on the solid stationary phase.
Adsorption
Not widely used in the lab because of technical problems with the preparation of stationary phase that has homogenous distribution of absorption sites
Adsorption
The molecules that are most soluble in the mobile phase move fastest; the least soluble move slowest.
Adsorption
can be acidic polar, basic polar or nonpolar
Stationary phase
can be a single solvent or mixture of two or more solvents, depending on the analytes to be desorbed
Mobile phase
Also referred to as liquid–liquid chromatography. Separation of solute is based on relative solubility in an organic (nonpolar) solvent and an aqueous (polar) solvent.
Partition
o Applicable to any substance that may be distributed between two liquid phases but works best with nonionic compound
o In its simplest form, partition (extraction) is performed in a separatory funnel.
Partition
In Partition, ______ molecules remain in the aqueous solvent; __________ molecules are extracted in the organic solvent.
Polar; nonpolar
uses pseudo-liquid stationary phases that are chemically bonded to the support or high-molecular-weight polymers that are insoluble in the mobile phase
Modern partition chromatography
o Variation of liquid–solid chromatography, is used to separate solute molecules on the basis of size and shape.
o A sample containing different-sized molecules moves down the column dissolved in the mobile solvent
Steric Exclusion
enter the pores in the packing and are momentarily trapped
Small molecules
excluded from the small pores and so move quickly between the particles
Large molecules
partially restricted from entering the pores and therefore, move through the column at and intermediate rate
Intermediate-sized molecules
Uses hydrophilic beads of cross-linked dextran, polyacrylamide or agarose → forms gel when soaked in water
Gel filtration
similar separation process using hydrophobic gel beads of polystyrene with a nonaqueous mobile phase
Gel permeation chromatography
o Solute mixtures are separated by virtue of the magnitude and charge of ionic species
Ion-Exchange Chromatography
o Anion-exchange resins are made with exchangeable hydroxyl ions such as the
diethylamine functional group
o Stationary phase is a resin, consisting of large polymers of substituted benzene, silicates, or cellulose derivatives, with charged functional groups
o used to remove interfering substances from a solution, to concentrate dilute ion solutions, and to separate mixtures of charged molecules, such as amino acid
Ion-exchange chromatography
variant of column chromatography. A thin layer of sorbent, such as alumina, silica gel, cellulose, or cross-linked dextran, is uniformly coated on a glass or plastic plate
Thin Layer Chromatography
- Mobile phase (solvent) is usually placed in a closed container until the atmosphere is saturated with solvent vapor
- The solvent migrates up the thin layer by capillary action, dissolving and carrying sample molecules.
Thin Layer Chromatography
- most commonly used as a semiquantitative screening test
- Plates prepared with uniform sorbent thickness, finer particles, and new solvent systems have resulted in the technique of high-performance thin layer chromatography (HPTLC)
Thin Layer Chromatography
