HPLC columns and classification MCQs With Answer

Introduction: This quiz collection on HPLC columns and their classification is designed specifically for M.Pharm students studying Advanced Instrumental Analysis (MPA 201T). It covers essential concepts such as stationary phase chemistries (C18, phenyl, cyano), particle technologies (fully porous, superficially porous/core–shell, monolithic), pore sizes, end-capping, speciality columns (ion-exchange, size-exclusion, HILIC, chiral), column dimensions and pressure considerations. Each question focuses on practical understanding and selection criteria used in method development and troubleshooting. Practising these MCQs will strengthen your grasp of column choice, chromatographic behavior, and technical parameters required for robust, high-resolution separations in pharmaceutical analysis.

Q1. Which stationary phase is most commonly used in reversed-phase HPLC?

• Octadecylsilane (C18)
• Silica gel (unmodified)
• Amino propyl
• Diol

Correct Answer: Octadecylsilane (C18)

Q2. What is the primary purpose of end-capping silica-based bonded phases?

• To reduce residual silanol activity
• To increase the pore size
• To act as a secondary ion-exchange site
• To increase carbon load dramatically

Correct Answer: To reduce residual silanol activity

Q3. Which advantage is associated with core–shell (superficially porous) particles compared to fully porous particles?

• Reduced mass transfer resistance and improved efficiency at comparable backpressure
• Much larger pore sizes suitable only for proteins
• Complete elimination of surface silanols
• They always produce lower retention for nonpolar compounds

Correct Answer: Reduced mass transfer resistance and improved efficiency at comparable backpressure

Q4. Selectivity in HPLC separations is most strongly influenced by which factor?

• Stationary phase chemistry and its interaction mechanism with analytes
• Particle size of the packing alone
• Column length by itself
• Mobile phase flow rate only

Correct Answer: Stationary phase chemistry and its interaction mechanism with analytes

Q5. Which of the following is NOT classified as an ion-exchange stationary phase?

• Sulfonic acid (strong cation exchanger)
• Quaternary ammonium (strong anion exchanger)
• Octadecyl (C18) bonded silica
• Carboxylate (weak cation exchanger)

Correct Answer: Octadecyl (C18) bonded silica

Q6. For analysis of peptides and small proteins, which pore size of silica packing is commonly recommended?

• 300 Å
• 30 Å
• 5 Å
• 10 Å

Correct Answer: 300 Å

Q7. What is a primary benefit of monolithic columns compared with conventional packed-particle columns?

• High permeability allowing higher flow rates with lower backpressure
• They provide stronger ion-exchange capacity by default
• They have no surface chemistry variability
• They require much larger sample volumes always

Correct Answer: High permeability allowing higher flow rates with lower backpressure

Q8. Which bonded phase gives enhanced π–π interactions useful for aromatic compounds?

• Phenyl-bonded phase
• C8 (octyl)
• Cyano (CN)
• Diol

Correct Answer: Phenyl-bonded phase

Q9. Compared to sub-2 µm fully porous particles, a 2.7 µm core–shell column typically provides which practical advantage?

• Similar efficiency at lower backpressure
• Higher chemical inertness at extreme pH
• Better compatibility with gel-permeation separations
• Wider pore size distribution for large proteins

Correct Answer: Similar efficiency at lower backpressure

Q10. What does the column efficiency parameter ‘N’ represent?

• Number of theoretical plates
• Plate height
• Column length in meters
• Retention factor

Correct Answer: Number of theoretical plates

Q11. Which stationary phase material is commonly used for size-exclusion chromatography of synthetic polymers?

• Cross-linked polystyrene–divinylbenzene
• Octadecylsilane (C18)
• Quaternary ammonium silica
• Phenyl-bonded silica

Correct Answer: Cross-linked polystyrene–divinylbenzene

Q12. How does increasing the carbon load of a reversed-phase column affect chromatographic behavior?

• Increases hydrophobic retention (longer retention for nonpolar analytes)
• Reduces retention for nonpolar analytes
• Has no effect on selectivity or retention
• Always increases peak broadening drastically

Correct Answer: Increases hydrophobic retention (longer retention for nonpolar analytes)

Q13. Which column type is most appropriate when extreme pH stability (pH 1–13) is required?

• Polymeric (e.g., polystyrene/divinylbenzene) columns
• Unmodified silica columns
• Standard bonded silica C18 columns without special treatment
• Typical silica-based HILIC columns

Correct Answer: Polymeric (e.g., polystyrene/divinylbenzene) columns

Q14. HILIC stationary phases are best suited to retain which class of analytes?

• Highly polar small molecules and polar metabolites
• Large nonpolar lipids only
• Proteins above 50 kDa exclusively
• Volatile organics used in gas chromatography

Correct Answer: Highly polar small molecules and polar metabolites

Q15. Which parameter most strongly influences column backpressure in packed HPLC columns?

• Particle size of the packing material
• Stationary phase chemistry only
• Detector type
• Sample concentration

Correct Answer: Particle size of the packing material

Q16. The retention factor (k’) is defined by which equation?

• k’ = (tR − t0) / t0
• k’ = t0 / tR
• k’ = tR × t0
• k’ = tR − t0

Correct Answer: k’ = (tR − t0) / t0

Q17. What is a principal analytical advantage of superficially porous (core–shell) particles?

• They give high efficiency with reduced diffusion path lengths, improving speed and resolution
• They eliminate the need for mobile phase optimization entirely
• They are only useful for ion-exchange separations
• They increase column void volume substantially

Correct Answer: They give high efficiency with reduced diffusion path lengths, improving speed and resolution

Q18. Which class of stationary phases is most commonly used for chiral separations in HPLC?

• Polysaccharide derivatives (e.g., cellulose or amylose derivatives)
• Pure unmodified silica gel
• Strong cation exchangers
• C8 reversed-phase only

Correct Answer: Polysaccharide derivatives (e.g., cellulose or amylose derivatives)

Q19. What is the primary purpose of using a guard column ahead of the analytical column?

• To protect the analytical column from particulates and strongly retained contaminants
• To increase theoretical plate count directly
• To act as a detector replacement
• To change the mobile phase composition dynamically

Correct Answer: To protect the analytical column from particulates and strongly retained contaminants

Q20. Which of the following is a common classification of HPLC columns based on their physical form?

• Packed bed (particles), monolithic, and open tubular
• Only by brand name and manufacturer
• Only by detector compatibility
• Only by the color of the column housing

Correct Answer: Packed bed (particles), monolithic, and open tubular

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