Ion exchange chromatography is a core separation technique in pharmaceutical analysis that separates molecules by charge using charged stationary phases (resins). In B. Pharm labs it’s essential for protein and peptide purification, drug impurity profiling, formulation characterization, and quality control. Key concepts include stationary phase chemistry (strong vs weak exchangers), functional groups (sulfonic, carboxylic, quaternary ammonium), resin backbone, exchange capacity, selectivity, pH and ionic strength effects, elution strategies (salt or pH gradients), gradient design, detection (conductivity), resin regeneration, and scale-up principles. A strong conceptual and practical grasp helps in method development and troubleshooting. Now let’s test your knowledge with 30 MCQs on this topic.
Q1. What is the fundamental separation principle of ion exchange chromatography?
- Size exclusion based on molecular diameter
- Hydrophobic interactions with the stationary phase
- Electrostatic interactions between charged analytes and charged stationary phase
- Specific affinity between ligand and target molecule
Correct Answer: Electrostatic interactions between charged analytes and charged stationary phase
Q2. Which statement correctly classifies ion exchange resins?
- Cation exchangers bind negatively charged analytes
- Anion exchangers bind positively charged analytes
- Cation exchangers have negatively charged functional groups and bind cations
- All ion exchangers are neutral and separate by size
Correct Answer: Cation exchangers have negatively charged functional groups and bind cations
Q3. Which functional group is typical of a strong cation exchanger?
- Carboxylate (-COO–)
- Sulfonic acid (-SO3–)
- Tertiary amine (-NR3)
- Hydroxyl (-OH)
Correct Answer: Sulfonic acid (-SO3–)
Q4. What is a common synthetic backbone material for high-stability ion exchange resins?
- Cellulose
- Agarose
- Polystyrene-divinylbenzene (PS-DVB)
- Silica gel without modification
Correct Answer: Polystyrene-divinylbenzene (PS-DVB)
Q5. How does pH primarily affect retention on weak ion exchangers?
- It changes the mobile phase viscosity only
- It alters ionization state of analyte and resin functional groups, affecting retention
- It affects only temperature of the column
- pH has no effect on weak exchangers
Correct Answer: It alters ionization state of analyte and resin functional groups, affecting retention
Q6. How is the total exchange capacity of an ion exchange resin defined?
- The flow rate at which the resin operates optimally
- Total amount of exchangeable ions per unit mass of dry resin (meq/g)
- Volume of mobile phase consumed per run
- Number of theoretical plates in the column
Correct Answer: Total amount of exchangeable ions per unit mass of dry resin (meq/g)
Q7. What does the selectivity coefficient describe in ion exchange?
- The temperature dependence of retention time
- The resin’s physical porosity
- The relative preference of the resin for one ion over another
- The flow resistance of the packed bed
Correct Answer: The relative preference of the resin for one ion over another
Q8. Which elution strategy increases ionic strength to displace bound ions?
- Isocratic elution with pure water
- Salt gradient elution (increasing counter-ion concentration)
- Temperature ramping
- Decreasing mobile phase ionic strength
Correct Answer: Salt gradient elution (increasing counter-ion concentration)
Q9. What is Donnan exclusion in the context of ion exchange resins?
- Preferential exclusion of co-ions (same charge as the fixed resin charge) from resin pores
- Complete neutralization of resin charge at any pH
- Physical exclusion of large molecules by pore size
- Hydrophobic shielding of charged sites
Correct Answer: Preferential exclusion of co-ions (same charge as the fixed resin charge) from resin pores
Q10. Which functionalized resin is an example of a strong cation exchanger used in protein separations?
- Carboxymethyl (CM)
- Sulfopropyl (SP)
- Diethylaminoethyl (DEAE)
- Hydroxylated silica
Correct Answer: Sulfopropyl (SP)
Q11. Which functional group is characteristic of a strong anion exchanger?
- Quaternary ammonium (-NR4+)
- Carboxylate (-COO–)
- Sulfonate (-SO3–)
- Phenyl ring
Correct Answer: Quaternary ammonium (-NR4+)
Q12. Which detector is commonly used for measuring ionic species eluting from an ion exchange column?
- UV-Vis at 280 nm only
- Fluorescence detector
- Conductivity detector
- Refractive index detector
Correct Answer: Conductivity detector
Q13. How does a high salt concentration in the sample solvent affect ion exchange binding?
- It enhances binding by reducing mobile phase ionic strength
- It reduces binding due to competition from sample ions
- It increases column efficiency automatically
- It has no effect on ion exchange binding
Correct Answer: It reduces binding due to competition from sample ions
Q14. In a cation exchange process, which ion typically serves as the counter-ion in the mobile phase?
- Chloride (Cl–)
- Sodium (Na+)
- Sulfate (SO4 2–)
- Hydroxide (OH–)
Correct Answer: Sodium (Na+)
Q15. At pH below its isoelectric point (pI), a protein will have which net charge and bind to which exchanger?
- Net negative; binds to anion exchanger
- Net positive; binds to cation exchanger
- Neutral; no binding to any exchanger
- Net negative; binds to cation exchanger
Correct Answer: Net positive; binds to cation exchanger
Q16. How is the capacity factor (k’) defined in chromatography?
- k’ = t0 / tR
- k’ = (tR – t0) / t0
- k’ = column length × flow rate
- k’ = mobile phase viscosity / temperature
Correct Answer: k’ = (tR – t0) / t0
Q17. Which action typically improves chromatographic resolution on an ion exchange column?
- Reducing the number of theoretical plates
- Decreasing column efficiency
- Increasing column efficiency (more theoretical plates)
- Using an irrelevant organic solvent
Correct Answer: Increasing column efficiency (more theoretical plates)
Q18. Which of the following is a common pharmaceutical application of ion exchange chromatography?
- Measuring vapor pressure of solvents
- Protein and peptide purification based on charge
- Measuring lipid partition coefficients only
- Assessing tablet hardness
Correct Answer: Protein and peptide purification based on charge
Q19. Which reagent is commonly used to regenerate a cation exchange resin to the H+ form?
- Sodium hydroxide (NaOH)
- Hydrochloric acid (HCl)
- Sodium chloride (NaCl)
- Ethanol
Correct Answer: Hydrochloric acid (HCl)
Q20. Which statement distinguishes strong ion exchangers from weak ion exchangers?
- Strong exchangers lose charge near neutral pH
- Strong exchangers maintain ionization (fixed charge) over a wide pH range
- Weak exchangers are always better for high pH
- Strong exchangers are only used for organic solvents
Correct Answer: Strong exchangers maintain ionization (fixed charge) over a wide pH range
Q21. What is the principle of displacement ion exchange chromatography?
- Using temperature to displace analytes from resin
- Introducing a displacer that has stronger affinity, continuously pushing analytes off the resin into discrete zones
- Using decreasing salt to elute bound analytes
- Partitioning analytes by hydrophobicity
Correct Answer: Introducing a displacer that has stronger affinity, continuously pushing analytes off the resin into discrete zones
Q22. In a cation exchange separation of Na+ and Ca2+, which ion typically elutes first under identical conditions?
- Ca2+
- Na+
- They coelute exactly
- Depends only on column temperature
Correct Answer: Na+
Q23. Which parameter primarily determines the order of elution in ion exchange chromatography?
- Column color
- Selectivity (affinity differences between ions)
- Pump brand
- Ambient humidity
Correct Answer: Selectivity (affinity differences between ions)
Q24. What is an advantage of gradient elution in ion exchange chromatography?
- It eliminates the need for buffers
- It shortens analysis time and helps elute strongly retained components
- It always reduces resolution
- It only works for neutral compounds
Correct Answer: It shortens analysis time and helps elute strongly retained components
Q25. How does resin pore size influence separation of large biomolecules?
- It only affects color of the eluate
- Pore size affects accessibility of binding sites and mass transfer, influencing resolution for large biomolecules
- Pore size is irrelevant for proteins
- Smaller pores always improve binding of large proteins
Correct Answer: Pore size affects accessibility of binding sites and mass transfer, influencing resolution for large biomolecules
Q26. What is the effect of decreasing particle size of the ion exchange resin?
- Decreases column backpressure
- Decreases column efficiency
- Increases efficiency but increases backpressure
- Makes the resin hydrophobic
Correct Answer: Increases efficiency but increases backpressure
Q27. How is the distribution coefficient (Kd) in ion exchange typically defined?
- Kd = volume of mobile phase / column length
- Kd = (amount of ion bound per mass of resin) / (equilibrium concentration in solution)
- Kd = flow rate × column diameter
- Kd = mobile phase pH × temperature
Correct Answer: Kd = (amount of ion bound per mass of resin) / (equilibrium concentration in solution)
Q28. What is the purpose of conductivity suppression in ion chromatography?
- To increase background conductivity for better signal
- To reduce background conductivity and enhance sensitivity for ionic analytes
- To make the mobile phase fluorescent
- To sterilize the column
Correct Answer: To reduce background conductivity and enhance sensitivity for ionic analytes
Q29. Which mobile phase component is generally NOT used in routine aqueous ion exchange chromatography?
- Buffered salt solutions
- Organic modifiers like low-percent methanol or acetonitrile
- High concentrations of nonpolar solvent like hexane
- pH-adjusting acids or bases
Correct Answer: High concentrations of nonpolar solvent like hexane
Q30. During scale-up of an ion exchange method from analytical to preparative column, which parameter should typically be maintained to preserve chromatographic behavior?
- Total column volume only
- Linear velocity (flow velocity through the column)
- Absolute flow rate without regard to column cross-section
- Only the column packing time
Correct Answer: Linear velocity (flow velocity through the column)
I am a Registered Pharmacist under the Pharmacy Act, 1948, and the founder of PharmacyFreak.com. I hold a Bachelor of Pharmacy degree from Rungta College of Pharmaceutical Science and Research. With a strong academic foundation and practical knowledge, I am committed to providing accurate, easy-to-understand content to support pharmacy students and professionals. My aim is to make complex pharmaceutical concepts accessible and useful for real-world application.
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