Chapter 1 - Introduction Flashcards
What is liquid chromatography?
a physical separation technique conducted between two phases - a solid phase and a liquid phase.
Chromatography meaning
“color writing”
HPLC advantages
- Applicable to diverse analytes
- Precise and highly reproducible quantitative analysis
- HPLC coupled with MS
- High separation power with sensitive detection
HPLC perceived limitations
- Lack of an ideal universal detector
- Less separation efficiency than capillary gas chromatography
- Still arduous for regulatory or quality control testing
HPLC is amenable to _____ % of all existing compounds.
60 to 80
GC is amenable for ______ % of all existing compounds.
15
Primary separation modes of HPLC
- Normal-Phase Chromatography (NPC)
- Reversed-Phase Chromatography (RPC)
- Ion-Exchange Chromatography (IEC)
- Size-Exclusion Chromatography (SEC)
NPC is particularly useful for the separation of
nonpolar compounds and isomers.
NPC uses a _______ stationary phase and a _______ mobile phase.
polar; nonpolar
One significant disadvantage of NPC is
easy contamination of the polar surfaces by highly retained sample components.
NPC is primarily used in
chiral separations and preparative applications.
RPC uses a ______ stationary phase and a ______ mobile phase.
nonpolar; polar
In RPC, polar analytes elute
first.
RPC typically uses a mixture of
methanol or acetonitrile with water.
The mechanism of separation in RPC is mainly attributed to
hydrophobic interaction.
RPC is used in approximately what percent of all HPLC analyses?
70%
In ion-exchange chromatography, the separation mode is based on
the exchange of ionic analytes with the counter ions of the ionic groups attached to the solid support.
In IEC, to force the migration of analytes, mobile phases consist of
buffers, often with increasing ionic strength (e.g. a higher concentration of NaCl)
Size-exclusion chromatography is based solely on
the analyte’s molecular size.
The goal of most HPLC analysis is to
separate analytes from other components in the sample for accurate quantitation.
Baseline resolution is usually not needed for MS detection since
specific signals can be customized for each ion according to its molecular weight.
