Applications of electrophoresis in drug analysis MCQs With Answer

Introduction

Electrophoresis is a vital analytical tool in drug analysis, widely used in pharmaceutical quality control, impurity profiling, formulation studies and forensic investigations. B.Pharm students should master capillary electrophoresis (CE), gel techniques (SDS-PAGE, native PAGE, agarose), isoelectric focusing (IEF) and hyphenated methods like CE-MS for the separation and identification of small molecules, peptides and protein therapeutics. Key concepts include electroosmotic flow, charge-to-size separation, MEKC for neutral drugs, buffer selection, sample stacking, detection modes (UV, LIF, MS) and method validation (LOD/LOQ, specificity, precision). Practical interpretation of electropherograms and impurity/glycoform analysis is essential for regulatory submissions. Now let’s test your knowledge with 30 MCQs on this topic.

Q1. What is the fundamental principle of electrophoresis used in drug analysis?

• The movement of neutral molecules by diffusion through a gel
• The movement of charged particles under an applied electric field through a medium
• The separation of compounds solely by molecular weight in solution
• The chemical reaction between analyte and buffer components

Correct Answer: The movement of charged particles under an applied electric field through a medium

Q2. Electrophoretic mobility of an ion in capillary electrophoresis is primarily determined by which factor?

• The absolute molecular weight only
• The charge-to-size (charge/mass or charge/hydrodynamic radius) ratio
• The buffer color and viscosity
• The detector wavelength used

Correct Answer: The charge-to-size (charge/mass or charge/hydrodynamic radius) ratio

Q3. Which CE mode is most appropriate for separating molecules solely based on their isoelectric point?

• Capillary zone electrophoresis (CZE)
• Micellar electrokinetic chromatography (MEKC)
• Isoelectric focusing (IEF)
• SDS-capillary gel electrophoresis

Correct Answer: Isoelectric focusing (IEF)

Q4. What is the main purpose of SDS in SDS-PAGE when analyzing protein drugs?

• To provide a pH gradient for isoelectric focusing
• To denature proteins and impart uniform negative charge for separation by molecular weight
• To act as a fluorescent label for detection
• To stabilize tertiary protein structure during electrophoresis

Correct Answer: To denature proteins and impart uniform negative charge for separation by molecular weight

Q5. What does electroosmotic flow (EOF) in capillary electrophoresis refer to?

• The chemical reaction between analyte and capillary wall
• The bulk movement of the background electrolyte toward an electrode due to charged capillary walls
• The diffusion-limited transport of neutral analytes only
• The optical detection response of the buffer

Correct Answer: The bulk movement of the background electrolyte toward an electrode due to charged capillary walls

Q6. Which electrophoretic technique is especially useful for separating neutral drug molecules?

• Isoelectric focusing (IEF)
• SDS-PAGE
• Micellar electrokinetic chromatography (MEKC)
• Agarose gel electrophoresis for nucleic acids

Correct Answer: Micellar electrokinetic chromatography (MEKC)

Q7. What is the main advantage of coupling capillary electrophoresis with mass spectrometry (CE-MS) in drug analysis?

• It eliminates the need for any buffer or separation
• High-efficiency separation combined with molecular identification and structural information from MS
• It increases sample volume requirements drastically
• It prevents Joule heating completely

Correct Answer: High-efficiency separation combined with molecular identification and structural information from MS

Q8. What is the purpose of stacking techniques (sample stacking) in CE for pharmaceutical analysis?

• To dilute the sample to avoid detector saturation
• To concentrate analytes into narrow zones to improve sensitivity and LOD
• To alter the pI of proteins prior to IEF
• To chemically derivatize analytes inside the capillary

Correct Answer: To concentrate analytes into narrow zones to improve sensitivity and LOD

Q9. Which detection technique generally offers the highest sensitivity for trace fluorescently labeled drug analytes in CE?

• UV-Visible absorbance detection
• Laser-induced fluorescence (LIF)
• Refractive index detection
• Conductivity detection without labeling

Correct Answer: Laser-induced fluorescence (LIF)

Q10. In an uncoated fused silica capillary at typical pharmaceutical pH (around neutral), what is the usual direction of electroosmotic flow?

• Toward the cathode (negative electrode)
• Toward the anode (positive electrode)
• There is no electroosmotic flow
• It alternates direction periodically during run

Correct Answer: Toward the cathode (negative electrode)

Q11. For impurity profiling of protein therapeutics, electrophoresis can effectively separate which of the following variants?

• Chromatographic retention time only
• Charge variants and glycoforms (isoforms) of the protein
• Only DNA contaminants
• Volatile organic impurities

Correct Answer: Charge variants and glycoforms (isoforms) of the protein

Q12. Which electrophoretic technique is most commonly used for sizing and visualizing DNA fragments in drug formulation or forensic samples?

• SDS-PAGE for proteins
• Agarose gel electrophoresis
• CE with only conductivity detection
• Isoelectric focusing (IEF)

Correct Answer: Agarose gel electrophoresis

Q13. Which additive is commonly used in CE as a chiral selector to achieve enantiomeric separation of chiral drugs?

• Sodium dodecyl sulfate (SDS) with no chiral properties
• Cyclodextrins (e.g., methyl-β-cyclodextrin)
• Acetic acid only
• Silver nitrate

Correct Answer: Cyclodextrins (e.g., methyl-β-cyclodextrin)

Q14. Which solvent system or CE variant is most suitable for analyzing poorly water-soluble small-molecule drugs?

• Standard aqueous CE only
• Nonaqueous capillary electrophoresis (NACE)
• Agarose gel electrophoresis
• IEF using ampholytes in water

Correct Answer: Nonaqueous capillary electrophoresis (NACE)

Q15. Which set of validation parameters is essential when establishing an electrophoretic method for quantitative drug impurity analysis?

• Color, smell and taste
• Specificity, precision, accuracy, linearity, LOD/LOQ
• Only migration time reproducibility
• Only buffer pH and ionic strength

Correct Answer: Specificity, precision, accuracy, linearity, LOD/LOQ

Q16. Two-dimensional electrophoresis commonly used in proteomics combines which two separation principles?

• Agarose electrophoresis followed by MEKC
• Isoelectric focusing (IEF) in first dimension and SDS-PAGE in second dimension
• SDS-PAGE twice with different voltages
• Capillary zone electrophoresis followed by agarose gel

Correct Answer: Isoelectric focusing (IEF) in first dimension and SDS-PAGE in second dimension

Q17. Which modification to capillary inner surface is typically used to reduce protein adsorption and control EOF?

• Leaving the capillary bare and unconditioned
• Dynamic or permanent coating with neutral or charged polymers
• Increasing applied voltage only
• Adding salts to increase ionic strength beyond normal limits

Correct Answer: Dynamic or permanent coating with neutral or charged polymers

Q18. What are the common sample injection methods in capillary electrophoresis?

• Only manual pipetting into the detector
• Hydrodynamic (pressure or vacuum) and electrokinetic injection
• Direct syringe infusion into MS without separation
• Immersion of capillary in sample for 24 hours

Correct Answer: Hydrodynamic (pressure or vacuum) and electrokinetic injection

Q19. Which statement best describes capillary zone electrophoresis (CZE)?

• A separation mode that uses micelles as pseudostationary phase
• A free-solution separation based on differences in charge-to-size ratio without micelles
• Equivalent to SDS-PAGE for proteins
• An electrophoretic method that requires a gel matrix inside the capillary

Correct Answer: A free-solution separation based on differences in charge-to-size ratio without micelles

Q20. In micellar electrokinetic chromatography (MEKC), how are neutral analytes separated?

• By formation of covalent bonds with the capillary wall
• By differential partitioning between micellar pseudostationary phase and aqueous phase
• By differences in isoelectric point only
• By enzymatic cleavage inside the capillary

Correct Answer: By differential partitioning between micellar pseudostationary phase and aqueous phase

Q21. Why is temperature control important in capillary electrophoresis runs for pharmaceutical samples?

• Temperature has no effect on electrophoretic separations
• Because Joule heating influences viscosity, diffusion and separation efficiency
• Only to prevent evaporation of large sample volumes
• Only to protect the detector from overheating

Correct Answer: Because Joule heating influences viscosity, diffusion and separation efficiency

Q22. Which strategy reduces Joule heating and improves separation performance in CE?

• Increasing buffer ionic strength indefinitely
• Using lower ionic strength buffers, reduced applied current, and efficient capillary cooling
• Using wider bore capillaries at very high voltages without cooling
• Adding salts that increase conductivity dramatically

Correct Answer: Using lower ionic strength buffers, reduced applied current, and efficient capillary cooling

Q23. How does capillary inner diameter affect separation and detection in CE?

• Larger diameter always gives better efficiency and sensitivity
• Smaller diameter increases separation efficiency and reduces Joule heating but can lower detection sensitivity
• Diameter has no measurable effect
• Smaller diameter eliminates need for detectors

Correct Answer: Smaller diameter increases separation efficiency and reduces Joule heating but can lower detection sensitivity

Q24. Which staining method provides the highest sensitivity for visualizing low-abundance proteins on gels?

• Coomassie Brilliant Blue staining
• Silver staining
• Simple water rinse
• UV absorbance without stain

Correct Answer: Silver staining

Q25. For quantitative electrophoretic analysis of small molecules in formulations, which approach is commonly used for accurate quantitation?

• Qualitative visual band assessment only
• CE with UV detection and external calibration curve or internal standard
• Running gels without standards
• Measuring migration time only without peak area

Correct Answer: CE with UV detection and external calibration curve or internal standard

Q26. Electrophoretic methods are particularly useful in biosimilar characterization because they can detect which critical quality attributes?

• Only potency by bioassay
• Charge heterogeneity, aggregation, and glycosylation variants
• Only excipient composition
• Only ionic contaminants unrelated to the protein

Correct Answer: Charge heterogeneity, aggregation, and glycosylation variants

Q27. In forensic drug analysis, capillary electrophoresis is advantageous for amphetamine enantiomer separation primarily because:

• It requires large sample volumes and long run times
• CE can employ chiral selectors to separate enantiomers rapidly with small sample volumes
• Enantiomers cannot be separated by CE
• It destroys the sample, preventing further testing

Correct Answer: CE can employ chiral selectors to separate enantiomers rapidly with small sample volumes

Q28. How does buffer pH influence electrophoretic separation of ionizable drug molecules?

• pH does not affect ionization or migration
• It alters the ionization state of analytes, changing charge and electrophoretic mobility
• Only temperature matters, not pH
• It only affects the color of the buffer

Correct Answer: It alters the ionization state of analytes, changing charge and electrophoretic mobility

Q29. For detailed peptide mapping and identification of post-translational modifications in monoclonal antibodies, which electrophoretic-hyphenated approach is most informative?

• SDS-PAGE alone with Coomassie staining
• Capillary electrophoresis coupled to mass spectrometry (CE-MS)
• Simple agarose gel electrophoresis
• Paper chromatography only

Correct Answer: Capillary electrophoresis coupled to mass spectrometry (CE-MS)

Q30. Why are electrophoretic techniques considered suitable for stability-indicating methods in regulatory submissions?

• They cannot separate degradation products
• They can resolve parent drug from degradation products, charge variants and impurities, supporting specificity and stability assessment
• They are only qualitative and not accepted by regulators
• They always require radioactive labeling to work

Correct Answer: They can resolve parent drug from degradation products, charge variants and impurities, supporting specificity and stability assessment

G S Sachin

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.

Mail- Sachin@pharmacyfreak.com