FInal: Supercritical Fluid Chromatography & Capillary Electrophoresis Flashcards
What are the properties of a supercritical fluid?
- the viscosity is similar to that of a gas
- supercritical fluid can move through a capillary or packed column without high pressures needed in HPLC
What kind of gradient is used in supercritical fluid chromatography?
a gradient with increasing pressure in order to achieve increasing solvent density which therefore increases the eluent strength.
Why are relative eluent strengths of solvents in adsorption chromatography fairly indpendent of solute?
because they are both competing for adsorption sites therefore it doesn’t matter how strong the solvent is in comparison with the solute.
the density of a supercritical fluid is close to that of a (solid/liquid/gas).
liquid
is a supercritical fluid a good solvent or a bad solvent?
good solvent
supercritical fluids behaves more like the _______ mobile phase in _____ than the ______ mobile phase in ________.
liquid; HPLC
gaseous; GC
How do you recover supercritical fluid samples?
allow the solution to equilibriate with the atomosphere at relatively low temperature.
What is supercritical fluid chromatography?
a hybrid of gas and liquid chromatography, using supercritical fluid as its mobile phase.
What type of compunds can SFC handle?
- nonvolatile or thermally unstable compounds
- compunds with no spectroscopic or electrochemical properties
When would you choose to use SFC over GC?
when dealing with compounds that are nonvolatile or thermally unstable.
when would you choose to use SFC over LC?
when dealing with a compound with no spectroscopic or electrochemical properties.
In hydrophilic interaction chromatography (HILIC), why is eluet strength increased by increasing the fraction of water in the mobile phase?
since the stationary phase is polar, increasing the fraction of water in the mobile phase will allow it to compete better with the stationary phase therefore decresing the eluent time.
Why is high pressure needed in HPLC?
it increases the flow rate, which allows the eluates to come out faster and obtain more effective separation between peaks.
what is a bonded phase in liquid chromatography?
a liquid stationary phase is bonded to the solid support.
What are criteria for an adequate isocratic chromotographic separation?
adequate separation in a resonable time (0.5~20)
and Rs > 1.5
What are the general steps in developing an isocratic separation for reversed-phase chromatography with one organic solvent
1) Goal of analysis
2) Sample Preparation
3) Detector
4) wide gradient used to determine if isocratic or gradient separation is to be used.
what is electrophoresis?
a separation method based on the differential rate of migration ions in an applied dc electric field
Define Electrophoretic Migration.
The movement of ions is caused by the electric field , or the potential difference between two electrodes
Name the direction in which cations, anions, and neutral molecules migrate.
cations mgrate towards the cathode
anions migrate toward the anode
neutral molecules do not favor either
In electrophoretic migration, what does the rate at which the molecules move depend on?
charge-to-size ratios
the (smaller/larger) the ratio, the faster an ion migrates in the electric field.
larger
List all the variables for controlling electroosmotic flow.
(8 variables)
- Electric Field
- Ionic Strength
- Organic Modifiers
- Surfactant
- Temperature
- Buffer pH
- Covalent Coating
- Neutral Hydrophilic
EF, IS, OM, S, T, B, CC, NH
What is the result of changing the Electric Field variable in Electroosmotic Flow.
proportional change in EOF.
*joule heating may result*
What is the result of changing the Buffer pH variable in Electroosmotic Flow.
EOF decreases at low pH, increases at high pH.
What is the result of changing the Ionic Strength variable in Electroosmotic Flow.
Decreases the zeta potential between the cation layers and EOF with increasing buffer concentration
*high ionic strength means high current and Joule heating*
What is the result of changing the Organic Modifiers variable in Electroosmotic Flow.
increasing modifier decreases the zeta potential & EOF
*complex effects*
What is the result of changing the Surfactant variable in Electroosmotic Flow.
Adsorbs to capillary wall through hydrophobic or ionic interactions.
*Anionic surfactants increase EOF*
*Cationic surfactants decrease EOF*
What is the result of changing the Neutral Hydrophillic variable in Electroosmotic Flow.
Adsorbs to capillary wall via hydrophobic interactions.
*Decreases EOF by shielding surface charge, also increases viscosity*
What is the result of changing the Covalent Coating variable in Electroosmotic Flow.
chemically bonded to capillary wall
*Many things can happen*
What is the result of changing the Temperature variable in Electroosmotic Flow.
changes the viscosity
*easy to control*
What variable(s) of the Van Deemter equation apply to capillary electrophoresis. why don’t some of the variables apply?
Only the B term
A(multiple path) term does not apply becasue its an open tube
C (mass transfer) doesn’t apply because there is no stationary phase
Where is the detector placed in CE?
at the cathode
What is the difference between HPLC and CE?
- CE has a flat flow vs HPLC has parabolic flow
- CE has higher separation efficiency than HPLC
- CE is simpler to operate than HPLC
- HPLC has more mobile and stationary phase choices
- CE is limited to thin capillaries while HPLC has variety.
Name the two types of systems in CE.
- Continuous system
- Discontinuous system
Name the different methods for continuous system electrophoresis.
- CZE: Caillary Zone Electrophoresis
- CEC: Capillary ElectroChromatography
- MEKC: Micellar Electrokinetic Capillary Chromat.
- CGE: Capillary Gel Electrophoresis
- CIEF: Capillary Isoelectric Focusing
Name the method for discontinuous system electrophoresis.
CITP: Capillary Isotachophoresis
What are the applications of CE
- Analysis of carbohydrates
- Analysis of inorganic anions/metals ions
- DNA profiling
- Protein identification
Describe Lab-on-a-Chip.
liquid can now be moved by Electroosmosis or by pressure through micron-size channels etched into glass or plastic chips the size of a microscope slide.
what are the different substrate materials for Lab-on-a-Chip.
silica
polymer
Name the different types of detection methods for Lab-on-a-chip.
Fluorescence
Electrochemistry
Mass Spectrometry
Although similar to HPLC, what are the distinguishable parts in supercritical fluid chromatography
- A thermostatted column
- A restrictor or back-pressure device
what is the purpose of a thermostatted column oven in super critical fluid chrom.
to provide precise temperature control of the mobile phase
what is the purpose of a restrictor or back-pressure device in super critical fluid chrom.
to maintain the desired temperature in the column and to convert the eluent from a super critical fluid to a gas for transfer to the detector
What are the variables for super critical fluid chromatography?
- pressure
- temperature
What is the most widely used mobile phase for supercritical fluid chromatography?
CO2
Why is CO2 the most widely used mobile phase in supercritical fluid chromatography.
- low critical temperature and pressureso its easily achieved and maintained.
- great solvent for non-polar organic molecules
- no absorbance in UV range
- odorless and nontoxic
- inexpensive
What dectors are used for supercritical fluid chromatography.
can use standard GC or HPLC detectors
what is one major advantage of SFC over HPLC?
flame ionization dectector can be used.
What are some applications of supercritical fluid chromatography.
- natural products
- drugs(chiral separation)
- foods
- pesticides and herbicides
- surfactants
- fossil fuels
- polymers
- explosives and propellants
list separation effciency of SFC, LC, and GC in order or greatest to least.
GC > SFC > LC
What are the features of an ideal extraction for supercritical fluids.
- rapid, simple, and inexpensive
- quantitative recovery without loss or degration
- yields a sufficiently concentrated solution of the analyte
- very little laboratory waste.
What is the reality of extraction for supercritical fluids.
- requires longer time to achieve enough analyte
- concentration step in required
- solvent cost is high
- degration and atmospheric pollution may occur.
Name the components that make up the instrumentation for supercritical flluid extraction
- A fluid source (CO2)
- A syringe pump (>400 atm; 2ml/min)
- A control valve
- An exit valve
What is the purpose of the separator and suppressor in suppressed-ion chromatography. For cation chromatography, why is the suppressor an anion-exchange membrane?
separator column: separates ions by ion exchange
suppressor column: reduces the conductivity of eluent by exchanging the counterion
the suppressor is anion exchange membrane in order to exchange an ion for OH-
State the effects of increasing cross-linking on an ion-exchange column.
decreased swelling, increased exchange capacity and selectivity, but longer equilibration time.
what is electroosmosis?
a pumping action where the excess cations in the diffuse part of the double layer migrates toward the cathode.
State three different methods to reduce electroosmotic flow.
- Lowering the pH
- varying tho ionic strength
- Increase the Modifier
Explain how neutral molecules can be separated by micellar electrokinetic chromatography
neutral molecules spends some time in the negativley charged micelles that are moving upstream with some electrophorectic velocity, and each neutral molecule has a different partition coefficent between the solution and the micelles which then means they can be thought of as a type of chromatography, because molecules will elute at different times.
what are the two parts distingishable in super critical fluid chromatography?
- Thermostatted column oven
- A restrictor or back pressure device
What does a thermostatted column oven provide in the supercritical fluid chromatography instrument
provides precise temperature control of the mobile phase
What does a restrictor or back-pressure device provide in the supercritical fluid chromatography instrument
maintains the desired temperature in the column and converts the eluent froma supercritical fluid to a gas for transfer to the detector
what is electrophoretic mobility (µep)
the constant of proportionality between the speed of the ion and the electric field strength
What is the best method to control electroosmotic flow?
Change the buffer pH
for molecules of similar size, mobilty (increases/decreases) with charge.
increases
what is joule heating?
when the flow of ions in the capillary generates heat
true or false: electrophoresis transports anions in the opposite direction from electoosmosis.
true
at neutral or high pH, why does electroosmosis transport anions to the cathode?
because electroosmosis is usually faster than electrophoresis, and electroosmosis is always moving toward the cathode.
At low pH, electroosmosis is weak and anions may never reach the detector. If you want to separate anions at low pH, what must you do?
reverse the polarity to make the sample end negative and the detector positive
The higher the apparent mobility, the (longer/shorter) the migration time.
shorter
