Mixed 1 Flashcards
What is the retention factor (k) sometimes called?
A. Separation factor
B. Capacity factor
C. Selectivity factor
D. Efficiency factor
B. Capacity Factor
Also known as retention factor, it is a convenient descriptor that is dimensionless and independent of flow rate and column dimensions
What is the void volume (V0) also known as?
A. Dead volume
B. Peak volume
C. Flow volume
D. Stationary volume
A. Dead volume
Void volume is also known as dead volume, hold-up volume, and sometimes retention volume of non-retained component
What determines the selectivity (α) of a chromatographic system?
A. The mobile phase composition
B. The ratio of retention factors of two analytes
C. The number of theoretical plates
D. The flow rate
B. The ratio of retention factors of two analytes
Selectivity (α) is determined as the ratio of the retention factors of two analytes, or the ratio of the reduced retention times
Which parameter is used as a measure of chromatographic efficiency?
A. Retention time (tR)
B. Number of theoretical plates (N)
C. Void volume (V0)
D. Retention factor (k)
B. Number of theoretical plates
The number of theoretical plates (N) is used as a measure of chromatographic efficiency
Identify the term: The product of the analyte retention time and the mobile-phase flow rate.
Retention Volume (VR)
Retention volume is calculated as the product of the analyte retention time and the mobile-phase flow rate
Identify the term: The part of the total analyte retention time that the analyte actually spends in the mobile phase moving through the column.
Void time (t0)
Void time (t0) is the part of the total analyte retention time that the analyte spends in the mobile phase moving through the column
Identify the term: The ratio of the distance between two peaks to the average width of these peaks.
Resolution (Rs)
Resolution (Rs) is the ratio of the distance between two peaks to the average width of these peaks
Which of the following components would be eluted with a retention factor equal to zero?
A. Components positively retained on the column
B. Nonretained components
C. Components with maximum retention time
D. Components with minimum peak width
B. Nonretained components
What is the relationship between the number of theoretical plates (N) and the efficiency of the separation?
A. Directly proportional
B. Inversely proportional
C. No relationship
D. Exponentially related
A. Directly proportional
Which parameter is independent of the mobile phase flow rate and column dimensions?
A. Retention time (tR)
B. Retention factor (k)
C. Void volume (V0)
D. Resolution (Rs)
B. Retention factor (k)
Selectivity (α) is primarily dependent on which of the following factors?
A. The nature of the analytes
B. The flow rate of the mobile phase
C. The temperature of the column
D. The length of the column
A. The nature of analytes
Identify the term: A dimensionless chromatographic descriptor that indicates how well a column retains an analyte.
Retention factor (k)
Identify the term: The hypothetical division of a chromatographic column into smaller sections to measure efficiency.
Theoretical Plates
The _______ is the difference between the total retention time and the hold-up time.
Reduced retention time (t’R)
What is the significance of the number of theoretical plates (N) in chromatography?
A. It indicates the flow rate of the mobile phase
B. It measures the efficiency of separation
C. It determines the selectivity of the column
D. It indicates the void volume
B. It measures the efficiency of separation
How is resolution (Rs) in chromatography defined?
A. The distance between two peaks divided by the flow rate
B. The distance between two peaks divided by the average width of the peaks
C. The peak width at half-height divided by the retention time
D. The ratio of the retention factors of two analytes
B. The distance between two peaks divided by the average width of the peaks
Identify the term: The difference between the analyte retention volume and the void volume.
Reduced retention volume (V’R)
Which term describes the analyte’s equilibrium between the stationary and mobile phases in the column?
Plate Theory
Identify the separation mechanism where the stationary phase is a polymer with different pore sizes.
Mechanical entrapment
What is the primary interaction in Normal Phase Liquid Chromatography (NPLC)?
A. Hydrophobic interactions
B. Ionic interactions
C. Polar interactions
D. Mechanical entrapment
C. Polar Interactions
The principle of separation in Reversed Phase Liquid Chromatography (RPLC) is similar to _______.
solvent-solvent partitioning
In Ion-exchange Liquid Chromatography, what is the primary basis for separation?
A. Size differences
B. Charge densities
C. Polarity
D. Hydrophobicity
B. Charge densities
What term describes the characteristic of molecules based on their structure and electron charge distribution?
Polarity
_______ chromatography is used to separate molecules based on size using a polymer with different pore sizes.
Size Exclusion
Which type of chromatography involves a competition between the stationary phase and the mobile phase for different analytes?
A. Adsorption Chromatography
B. Partition Chromatography
C. Ion-exchange Chromatography
D. Size Exclusion Chromatography
A. Adsorption Chromatography
In Ion-exchange Liquid Chromatography, what is the primary basis for separation?
A. Size differences
B. Charge densities
C. Polarity
D. Hydrophobicity
B. Charge densities
Which type of chromatography involves a competition between the stationary phase and the mobile phase for different analytes?
A. Adsorption Chromatography
B. Partition Chromatography
C. Ion-exchange Chromatography
D. Size Exclusion Chromatography
A. Adsorption Chromatography
In Size Exclusion Chromatography (SEC), what is the basis for separation?
A. Polarity
B. Size
C. Charge
D. Hydrophobicity
B. Size
The number of theoretical plates (N) is a measure of _______ in chromatography.
efficiency
Which type of liquid chromatography uses a liquid stationary phase supported by a solid?
A. Normal Phase Liquid Chromatography
B. Reversed Phase Liquid Chromatography
C. Ion-exchange Liquid Chromatography
D. Size Exclusion Chromatography
B. Reversed Phase Liquid Chromatography
In which type of chromatography is the stationary phase an adsorbent?
A. Partition Chromatography
B. Ion-exchange Chromatography
C. Adsorption Chromatography
D. Size Exclusion Chromatography
C. Adsorption Chromatography
Which mode of chromatography separates molecules based on charge densities?
A. Normal Phase Liquid Chromatography
B. Reversed Phase Liquid Chromatography
C. Ion-exchange Liquid Chromatography
D. Size Exclusion Chromatography
C. Ion-exchange Liquid Chromatography
What is the principle of retention in Normal Phase Liquid Chromatography (NPLC)?
A) More polar solutes are preferentially retained or adsorbed on the polar stationary phase.
B) More non-polar solutes are preferentially retained or adsorbed on the non-polar stationary phase.
C) Both polar and non-polar solutes are equally retained.
D) None of the above.
A) More polar solutes are preferentially retained or adsorbed
Identify the most commonly used adsorbent material in NPLC.
A) Alumina
B) Titania
C) Zirconia
D) Silica
D) Silica
Fill in the blank: Silica is ______, causing extra strong retention of analytes with amino groups.
A) basic
B) acidic
C) neutral
D) amphiprotic
B) acidic
Which of the following statements about alumina, titania, and zirconia is correct?
A) They have basic functions, giving strong retention of analytes with acidic groups.
B) They are Lewis acids, retaining Lewis bases like ethers, ketones, and aromatics.
C) Titania- and zirconia-based materials have been used for high-temperature HPLC applications.
D) All of the above.
D) All of the above
Identify the stationary phases that offer different selectivity when bonded to silica.
A) Cyano and amino
B) Amino and diol
C) Cyano and diol
D) All of the above
D) All of the above
Which of the following solvents are considered weak solvents in adsorption chromatography?
A) Alcohols
B) Water
C) Acetonitrile
D) Alkanes
D) Alkanes
True or False: Alcohols, water, and acetonitrile bind to the silanol groups, deactivate silica, and prevent strongly tailing peaks.
True
Which of the following statements about gradient elution in NPLC is correct?
A) Gradient elution is not possible due to solvent de-mixing.
B) Gradient elution is preferred over isocratic elution.
C) Gradient elution is used to improve resolution.
D) None of the above.
A) Gradient elution is not possible due to solvent de-mixing.
Identify the application of NPLC where it is advantageous over Reversed-Phase Liquid Chromatography (RPLC).
A) Separation of very hydrophobic molecules
B) Separation of very hydrophilic molecules
C) Analysis of compounds prone to hydrolysis
D) All of the above
B) Separation of very hydrophilic molecules
Which of the following factors contribute to isomer selectivity in NPLC separations on silica columns? (Select all that apply)
A) Steric hindrance
B) Electron donation
C) Relative positions of polar groups within the solute molecule
D) Intramolecular hydrogen bonding of two polar groups
E) All of the above
E) All of the above
True or False: Gradient elution is possible in NPLC.
False. Due to the bound polar molecules, silica cannot be used for gradient elution because equilibration to starting conditions takes too long time.
Formation of ______ and ______ can cause problems when using lower boiling point solvents in NPLC.
A) bubbles, evaporation
B) precipitation, cloudiness
C) decomposition, precipitation
D) cloudiness, decomposition
A) bubbles, evaporation
Which of the following is NOT a disadvantage of NPLC?
A) It is not suitable for the analysis of a wide range of compounds.
B) Gradient elution is not possible due to solvent de-mixing.
C) Formation of bubbles and evaporation can occur with lower boiling point solvents.
D) It is difficult to control the polarity of the mobile phase.
D) It is difficult to control the polarity of the mobile phase.
Identify the correct statement about the retention behavior of polar and non-polar solutes in NPLC.
A) Polar solutes are more retained than non-polar solutes.
B) Non-polar solutes are more retained than polar solutes.
C) Both polar and non-polar solutes have similar retention.
D) None of the above.
A) Polar solutes are more retained than non-polar solutes.
Which of the following statements about the use of alcohols as mobile phase solvents in NPLC is correct?
A) Alcohols are weak solvents in adsorption chromatography.
B) Alcohols bind to the silanol groups, deactivating silica and preventing strongly tailing peaks.
C) Alcohols are preferred over alkanes for NPLC separations.
D) Both B and C are correct.
E) Both A and C are correct.
D) Both B and C are correct.
Identify what is not an application of NPLC.
A) Separation of very hydrophobic molecules
B) Separation of very hydrophilic molecules
C) Analysis of compounds prone to hydrolysis
D) Chiral separations
D) Chiral separations
True or False: The retention time of components can be variable in NPLC.
True
Fill in the blank: _______ and _______ have been used as high-temperature HPLC stationary phases due to their high stability, often coated with phosphates to reduce strong Lewis acid properties.
A) Silica, alumina
B) Alumina, zirconia
C) Titania, zirconia
D) Cyano, amino
C) Titania, zirconia
Which statement best describes the relationship between solute polarity and retention in NPLC?
A) More polar solutes are less retained than non-polar solutes.
B) More polar solutes are more retained than non-polar solutes.
C) Solute polarity does not affect retention in NPLC.
D) Both polar and non-polar solutes have similar retention.
B) More polar solutes are more retained than non-polar solutes.
Which of the following statements about the stationary phases used in NPLC is incorrect?
A) Silica is the most commonly used adsorbent material.
B) Alumina and titania have basic functions, retaining analytes with acidic groups.
C) Polar-bonded phases like cyano, amino, and diol offer different selectivity than silica.
D) Silica can be used for gradient elution in NPLC.
D) Silica can be used for gradient elution in NPLC.
Identify the isomers shown in Figure 9a.
A) cis and trans isomers of Doxepin
B) cis and trans isomers of Nortriptyline
C) cis and trans isomers of N-desmethyldoxepin
D) None of the above
A) cis and trans isomers of Doxepin
In Figure 11, which mobile phase composition (hexane-dichloromethane-2-propanol) provides better separation of the steroid isomers 3-6?
a.
A) 82:10:8 (v/v/v)
B) 84:10:6 (v/v/v)
C) Both compositions provide similar separation
D) None of the given compositions separate the isomers
B) 84:10:6 (v/v/v)
Which solvent is considered the weakest on silica?
A) Hexane
B) Dichloromethane
C) Isopropanol
D) Water
A) Hexane
What is the principle of separation in Size Exclusion Chromatography (SEC)?
A) It separates molecules based on their charge.
B) It separates molecules based on their hydrophobicity.
C) It separates molecules based on their molecular size.
D) It separates molecules based on their polarity.
C) It separates molecules based on their molecular size.
Identify the process that SEC separation relies on.
A) Adsorption of molecules on the packing material surface
B) Absence of interactions with the packing material surface
C) Ion-exchange between molecules and the packing material
D) Hydrophobic interactions between molecules and the packing material
B) Absence of interactions with the packing material surface
The two types of SEC discussed are ______ and ______.
A) Gel permeation chromatography (GPC), Gel filtration chromatography (GFC)
B) Ion-exchange chromatography, Affinity chromatography
C) Hydrophobic interaction chromatography, Reversed-phase chromatography
D) Adsorption chromatography, Partition chromatography
A) Gel permeation chromatography (GPC), Gel filtration chromatography (GFC)
Which of the following is NOT an application of SEC?
A) Determining the molecular weight distribution of polymers
B) Preparative fractionation of polymers
C) Purification of biological samples
D) Chiral separations
D) Chiral separations
Identify the mode of separation in SEC used to remove small molecules from a group of larger molecules.
A) High-resolution fractionation
B) Group separation
C) Ion-exchange separation
D) Reversed-phase separation
B) Group separation
Which of the following statements about group separation in SEC is correct?
A) It is often used in protein purification schemes for desalting and buffer exchange.
B) Small molecules such as excess salt are retained longer than larger molecules.
C) Sephadex G-10, G-25, and G-50 are not used for group separations.
D) None of the above.
A) It is often used in protein purification schemes for desalting and buffer exchange.
True or False: The goal of high-resolution fractionation in SEC is to separate multiple components in a sample based on their size differences.
True
Identify the stationary phase material used in Sephadex, a common SEC stationary phase.
A) Polyacrylamide
B) Styrene-divinyl benzene
C) Dextran
D) Agarose
C) Dextran
Which of the following statements about Sephadex LH-20 is correct?
A) It is a hydrophilic stationary phase material.
B) It is used for the fractionation of lipids, steroids, and other small molecules.
C) It swells only in water.
D) Both A and C are correct.
B) It is used for the fractionation of lipids, steroids, and other small molecules.
Sephadex LH-20 is a beaded, cross-linked dextran which has been hydroxypropylated to yield a chromatographic media with both hydrophilic and lipophilic character… designed for use in aqueous buffer systems, polar organic solvents, and aqueous solvent mixtures… fractionation of lipids, steroids, fatty acids, hormones, vitamins, and other small molecules.
Which of the following statements about Sephacryl is incorrect?
A) It is prepared by covalently cross-linking allyl dextran with N,N-methylene bisacrylamide.
B) It can be used in aqueous buffer systems with a pH range of 2-11.
C) Sephacryl S-200 and S-300 are useful for most proteins.
D) Sephacryl S-1000 fractionates small peptides and amino acids.
D) Sephacryl S-1000 fractionates small peptides and amino acids.
Sephacryl S-1000 fractionates restriction fragments of DNA, very large polysaccharides, proteoglycans, and membrane-bound vesicles op to 300-400 nm in diameter
What is an advantage of Superdex stationary phases?
A) They have high resolution and selectivity.
B) They have low non-specific interactions, permitting high recovery of biological material.
C) They combine the properties of cross-linked dextran and highly cross-linked agarose.
D) All of the above.
D) All of the above.
Superose stationary phases are based on highly cross-linked ______ particles and have ______ stability.
A) dextran, low
B) agarose, high
C) polyacrylamide, moderate
D) silica, high
B) agarose, high
Which of the following statements about Superose stationary phases is incorrect?
A) They allow the use of viscous eluents like 8 M urea at relatively high flow rates.
B) They exhibit negligible non-specific interactions with proteins at ionic strengths of 0.15 M to 1.5 M.
C) They show no hydrophobic interactions with any compounds.
D) Hydrophobic interactions can be an advantage for increasing resolution in some applications.
C) They show no hydrophobic interactions with any compounds.
Identify the stationary phase material used for the separation of synthetic (organic-soluble) polymers in Gel Permeation Chromatography (GPC).
A) Sephadex
B) Sephacryl
C) Highly cross-linked styrene-divinyl benzene
D) Superose
C) Highly cross-linked styrene-divinyl benzene
Which of the following stationary phases is NOT compatible with water-miscible organic solvents?
A) Highly cross-linked styrene-divinyl benzene
B) Bio-gel-P
C) Sephadex
D) Sephacryl
B) Bio-gel-P
What is the primary application of Enzacryl stationary phases?
A) Immobilization of enzymes
B) Separation of proteins
C) Separation of nucleic acids
D) Desalting of samples
A) Immobilization of enzymes
Which type of mobile phase is typically used in Size Exclusion Chromatography?
A) Non-polar organic solvents
B) Polar organic solvents
C) Aqueous buffer systems
D) Ion-pairing reagents
C) Aqueous buffer systems
True or False: The buffer composition generally does not influence the resolution in SEC unless it affects the shape or biological activity of the molecules.
True
Which of the following additives can be used in SEC mobile phases to maintain the solubility of proteins?
A) Urea and guanidine hydrochloride
B) Detergents
C) Both A and B
D) None of the above
C) Both A and B
What is the potential issue with using high concentrations of additives like urea or guanidine hydrochloride in SEC mobile phases?
A) They can increase the viscosity of the buffer, requiring lower flow rates.
B) They can cause precipitation of proteins.
C) They can deactivate the stationary phase.
D) They can increase non-specific interactions.
A) They can increase the viscosity of the buffer, requiring lower flow rates.
Based on the principle of SEC, which component would elute first from the column?
A) The smallest molecule in the sample
B) The largest molecule in the sample
C) All molecules elute simultaneously
D) The order of elution cannot be determined
B) The largest molecule in the sample
Fill in the blanks: In SEC, the separation process is based on the ability of sample molecules to ______ inside the pores of the packing material, which depends on the ______ of the analyte molecules and the pore size.
A) adsorb, charge
B) penetrate, relative size
C) ionize, hydrophobicity
D) partition, polarity
B) penetrate, relative size
Which of the following statements about the use of denaturing agents in SEC is incorrect?
A) Denaturing agents like urea or guanidine hydrochloride can be used for initial solubilization of the sample.
B) Denaturing agents can be used in SEC buffers to maintain solubility of proteins.
C) Denaturing agents should be avoided if denaturation of proteins is not desired.
D) Denaturing agents are useful for molecular weight determination of proteins.
C) Denaturing agents should be avoided if denaturation of proteins is not desired.
True or False: Extremes of pH, ionic strength, and the presence of denaturing agents or detergents can cause conformational changes, dissociation, or association of protein complexes in SEC.
True
What is the principle of separation in Ion Exchange Chromatography (IEX)?
A) It separates based on molecular size.
B) It separates based on hydrophobic interactions.
C) It separates based on ionic (or electrostatic) interactions between analytes and ionic functional groups on the stationary phase.
D) It separates based on partitioning between two immiscible phases.
C) It separates based on ionic (or electrostatic) interactions between analytes and ionic functional groups on the stationary phase.
Identify the two distinct mechanisms of separation in IEX.
A) Ion exchange and ion exclusion
B) Adsorption and desorption
C) Partitioning and ion exchange
D) Ion exchange and hydrophobic interactions
A) Ion exchange and ion exclusion
The two formats of ion exchangers are ______ and ______.
A) cation exchangers, anion exchangers
B) normal phase, reversed phase
C) hydrophobic, hydrophilic
D) size exclusion, ion exchange
A) cation exchangers, anion exchangers
Which of the following is NOT an application of IEX?
A) Separation of inorganic ions (ion chromatography)
B) Separation of biomolecules like proteins and oligonucleotides
C) Separation of carbohydrates
D) Chiral separations
D) Chiral separations
Identify the general component of an ion-exchange chromatograph that is used to introduce the sample onto the column.
A) High pressure pump
B) Injector
C) Column
D) Detector
B) Injector
