Exam 2 Flashcards
Normal-Phase Chromatography
Polarity of phases: Stationary Phase is Polar
Eluent Strength of solvent: Increase as solvent becomes more polar
Nature of Solutes: Polar
Nature of Solute interaction:
More soluble in mobile phase as polarity increases
Reversed-phase Chromatography
Polarity of phases: Stationary Phase is Non- Polar
Eluent Strength of solvent: Increase as solvent becomes less polar
Nature of Solutes: Non-Polar
Nature of Solute interaction:
More soluble in mobile phase as polarity decreases
Bonded reversed-phase
FM>2000, Soluble in octane, size 10nm
Molecular exclusion
FM>2000, Soluble in butanol, size 100nm
Bonded reversed-phase
FM>2000, Soluble in water, nonionic, size 10nm
Bonded normal-phase
FM<2000, Soluble in water, ion-paired
Ion-exchange
FM<2000, Soluble in water, ionic
A mixture of six nonpolar compounds was introduced to an HPLC with a C18 stationary phase for separation. How would the retention time and peak resolution for the mixture be affected if the solvent ratio was changed from 30% water: 70% acetonitrile to 15% water: 85% acetonitrile?
shorter retention time, lower peak resolution
The two components below are part of the mixture described above. Which will elute through this column first if the pH of the solution is 4.0?
CH3(CH2)5COOH—–CH3(CH2)5NH2
CH3(CH2)5NH2
Which of the following statements are true if the particle size in a packed column used in HPLC is decreased?
Increased resolution
Requires a higher operating pressure
Which of the following is a measure of kA?
(tA–tm)/tm
Which of the following is a measure of the resolution of A and B?
(tB–tA)/wavg
How might tm be determined? (Check all that apply).
from column volume and flow rate
from first disturbance of baseline
from first disturbance of baseline
a) Reversed phase column using 50% tetrahydrofuran/50% water.
b) Normal phase column using 50% hexane/50% diethyl ether.
a) Lower concentration of tetrahydrofuran to lower solvent strength.
b) a) Lower concentration of tetrahydrofuran to lower solvent strength.
b
Which of the following criteria meet the requirements to achieve and maintain an adequate isocratic HPLC separation?
- Operating pressure = 12 MPa
- k = 15
- Resolution ≥ 2
Gas Chromatography Detectors
- flame ionization detector
- thermal conductivity detector
- electron capture detector
Detectors for both Gas & Liquid Chromatography
mass spectrometer
Liquid Chromatography Detectors
- ultraviolet detectors
- charged aerosol dectector
Arrange the following solvents used in HPLC with ultraviolet detection from longest ultraviolet wavelength cutoff to shortest wavelength cutoff.
Longest- Acetone Toluene Dichloromethane Methanol Acetonitrile -Shortest
What is the ultraviolet wavelength cutoff for water?
190nm
It is the most common HPLC detector. It is useful for gradient elution with nonabsorbing solvents. Simple systems use lamps that produce discrete radiation, such as the 254-nm Hg vapor lamp, while more versatile systems use lamps that produce broadband radiation, such as xenon or tungsten lamps, and a monochromator to choose the wavelength of analysis.
Ultraviolet detector
With this detector, no peaks from the solvent are produced and it is compatible with gradient elution. The eluate is nebulized, forming an aerosol that is then passed through a drying tube where the solvent is evaporated, leaving analyte aerosol particles. The particles scatter light from a diode laser, generating the signal for the chromatogram. The detector response is related to the mass of the analyte, not its structure or molecular mass. The use of low-concentration buffers is required with this detector.
Evaporative light-scattering detector
A detector with virtually no selectivity that is not compatible with gradient elution. It uses filtered visible light as the irradiation source. The main appeal of this detector is its universal response to all solutes, even those with little absorption in the ultraviolet region of the spectrum.
Refractive Index Detector
The dynamic range of this sensitive detector covers 4-5 orders of magnitude in analyte concentration. The eluate is first nebulized in a stream of N2 gas then a positive charge is added to the aerosol particles as they pass over a charged Pt needle. Small ions are attracted to the charged plates of a small ion trap, while the larger aerosol particles are too large to be deflected and pass to a collector where they are detected by an electrometer.
Charged aerosol detector
Sort the HPLC detectors by compatibility with separation gradient.
Compatible with Gradient Elution:
- Ultraviolet Detector
- Evaporative Light-Scattering Detector
- Charged Aerosol Detector
NOT Compatible with Gradient Elution
- Electrochemical Detector
- Refractive Index Detector
HPLC columns that employ a liquid stationary phase, such as octadecyl (C18), covalently bound to a silica support material must be operated within a specific pH range. Which of the following pH ranges will allow a HPLC column with a stationary phase covalently bonded to an unmodified silica support to operate optimally?
3.0-6.5 & 2.9-4.5
It is packed or open tubular and is used to separate the analytes.
Column
It is maintained approximately 50 °C above the boiling point of the least volatile analyte.
Injector Port
It is maintained at a temperature slightly greater than the boiling point of the least volatile analyte.
Column Oven
Common ones include thermal conductivity, flame ionization, or electron capture.
Detector
In gas chromatography, what are the advantages of (a) temperature programming? (check all that apply.) b)pressure programming
a) Resolution of low-boiling solutes is maintained.& Retention times of high-boiling solutes are decreased.
b)
- Resolution of low-boiling solutes is maintained.
- Retention times of high-boiling solutes are decreased.
- Thermal degradation of samples is less likely.
- It isn't necessary to wait for a hot column to cool before injecting another sample.
In gas chromatography why do open tubular columns yield greater solute resolution than packed columns?
- Open tubular columns have a lower resistance to gas flow, so longer columns can be used without increasing solute retention times.
- Open tubular columns eliminate the multiple path term (A) from the van Deemter equation
In gas chromatography why can more rapid flow rates be used with H2 and He carrier gases than with N2, without sacrificing column efficiency?
Solutes diffuse more rapidly through H2 and He than through N2.
analyte can decompose above its boiling point, so the injector is not heated
On-column injection
analytes consist of less than 0.01% of sample
Split less Injection
analytes consist of more than 0.1% of sample
Split injection
Which of the following trapping types in splitless injection requires initially cooling the column temperature 150°C lower than the solute boiling points, and then raising the column temperature to vaporize solvent at the start of the column?
Cold Trapping
Which of the following trapping types in splitless injection removes solvent by setting the initial column temperature 40°C below the solvent boiling point to condense solvent at the front of the column?
Solvent Trapping
Carbon atoms bearing hydrogen atoms
Thermal Conductivity
Mass spectrometer
Flame ionization
Molecules with halogens, CN, NO2, conjugated C=O
Thermal Conductivity
Mass spectrometer
Electron capture
Aromatic and unsaturated compounds
Thermal Conductivity
Mass spectrometer
Photoionization
P and S and other elements selected by wavelength
Thermal Conductivity
Mass spectrometer
Flame photometric
P and N (and also hydrocarbons)
Thermal Conductivity
Mass spectrometer
Nitrogen-Phosphorus
S
Thermal Conductivity
Mass spectrometer
sulfur Chemiluminescence
Which of the following are benefits for choosing air from the atmosphere as a carrier gas in gas chromatography?
- does not require a bulky supply tank
- readily accessible
- Inexpensive
Which of the following carrier gases is used to decrease the loss of column efficiency at high flow rates, because it allows the solutes to diffuse faster from the center of the column to the wall?
H2
Which of the following molecules would a flame ionization detector (FID) be sensitive to after separation on a gas chromatography column? Choose all that apply.
3-methylpentane
I-pentane
methane
FIDs are sensitive to most hydrocarbons such as methane, i-pentane, and 3-methylpentane. FIDs are insensitive to nonhydrocarbons such as hydrogen, nitrogen monoxide, carbon dioxide, and hydrogen sulfide.
Thermal conductivity detectors in gas chromatography respond to changes in the thermal conductivity of the eluate. Due to this, ideally the difference in conductivities of the carrier gas and the solute should be as great as possible. The two carriers of choice for use in thermal conductivity detection are
He & H2
Sort the following parameters by whether they cause band broadening in gas-liquid chromatography.
- Very low flow rates
- low temperature
- large packing particles
- thick stationary phase layer
Sort the following parameters by whether they cause band separation in gas-liquid chromatography.
- small packing particles
- thin stationary phase layer
- rapid sample injection
Can be made of stainless steel or glass with a packed solid support of silanized silica
Packed Column
A column with a liquid stationary phase bonded to its wall
Wall-coated open tubular column
A column with a solid stationary phase coating the inside wall.
Porous-layer open tubular column
Has a solid support that is coated with a liquid stationary phase.
Support-coated open tubular column
Choose the statements that are true regarding open tubular and packed columns in gas chromatography. (Hint: there is more than one correct answer).
- For similar analysis times, open tubular columns provide better resolution than packed columns
- Packed columns have a greater sample capacity than open tubular columns.
- Packed columns give broader peaks with longer retention times.
Which method is used to reduce stationary phase degredation (bleeding) in gas chromatography?
Covalently bonding the stationary phase to the support material.
Choose the column performance chacteristics that can be used to indicate the degradation of the stationary phase in gas chromatography? Check all that apply.
- The retention factor of a known standard.
- The number of plates.
- Peak asymmetry.
A method for isolating compounds for analysis by gas chromatography that uses a stationary phase-coated fiber to extract compounds from air or liquid samples
Solid-Phase microextraction
Used to vaporize the analye after extraction in preparation for analysis by gas chromatography.
Thermal desorption tube
A sensitive extraction technique for isolating compounds from liquid samples by a magentic rod coated with a layer of stationary phase material.
Stir-bar sorptive extraction
A method of analyte condensation at the begining of the column to produce a narrow injection band.
Cold Trapping
Uses a chemical reaction to convert an analyte to a form more easily separated and detected
Derivatization
An extraction method with 100% efficiency used for removing volatile analytes from solid or liquid samples.
Purge & Trap
Extracting a portion of available analyte from a sample without the use of solvent
Solid-phase microextraction
Increasing the concentration of solute in a sample by the removal of solvent.
Preconcentration
Which of the following correctly describes solid-phase microextraction?
A method for isolating compounds for analysis by gas chromatography that uses a stationary phase coated fiber to extract compounds from gas or liquid samples
What is the purpose of cold trapping in gas chromatography?
To condense the analyte at the start of a gas chromatography column to insure a narrow injection band.
Why is stir-bar sorptive extraction a more sensitive extraction technique than solid-phase microextraction?
The volume of stationary phase that coats the stir bar in stir-bar sorptive extraction is significantly greater than the volume of stationary phase that coats the fiber in solid-phase microextraction.
A young researcher set out to develop a method for quantifying the polycyclic aromatic hydrocarbon (PAH) content in a contaminated soil sample by gas chromatography. During the initial stages of method development, the researcher was unable to separate anthracene and naphthalene with baseline resolution. From the options below, select all possible modifications the researcher could make to improve the resolution.
- Increase the stationary phase thickness
- Use a column with a smaller inner diameter
- Increase the column length
Isocratic elution
one solvent or one mixture of
solvents is used for a separation
Gradient elution
solvent makeup is changed
during a separation
Ideally, any detector should:
- be sensitive to low concentrations of all analytes
- Provide linear response
- not broaden the eluted peaks
Start with easiest adjustment, move to hardest
- Change solvent strength by varying fraction of solvent
- Change temperature
- Change pH (small steps!)
- Change to different solvent
- Change to different SP
dwell volume
Volume between where solvents are mixed and where
they enter the column
Gas-liquid partition chromatography
SP is
nonvolatile liquid bonded to inside of column or
to a fine solid support
Gas-solid adsorption chromatography
analyte
adsorbed directly on solid SP particles
Compared to packed, open tubular columns offer
- Higher resolution
- Shorter analysis time
- Greater sensitivity
- Lower sample capacity
SP can be lost due to
age, degradation at high temp,
evaporation, etc.
l Can lead to tailing if solutes bind to exposed SiOH groups
In order to prevent bleeding
SP is typically bonded
(covalently) to silica and covalently cross-linked to
itself
To monitor column performance
periodically
measure retention factor of a standard, plate number
and peak symmetry
l Changes indicate degradation of column 8
Compared to open tubular, packed columns have
- Greater sample capacity
- Broader peaks
- Longer retention times
- Less resolution
