Paper chromatography is a classical analytical technique that separates components by the principle of partition between a liquid mobile phase and the thin aqueous layer adsorbed on cellulose paper as the stationary phase. Widely used in pharmaceutical analysis, it employs development techniques such as ascending, descending, horizontal and radial chromatography; choice of solvent system, chamber saturation, sample application and paper type critically affect resolution and Rf values. Paper chromatography enables qualitative identification, impurity profiling, preparative separations and semi-quantitative estimation using UV, staining or densitometry. B. Pharm students should understand mechanism, development techniques, influencing factors, calculations and pharmaceutical applications. Now let’s test your knowledge with 30 MCQs on this topic.
Q1. Which fundamental mechanism primarily explains separation in paper chromatography?
- Adsorption onto silica particles in the paper.
- Partition between mobile solvent and aqueous layer on cellulose.
- Size-exclusion through paper fiber pores.
- Covalent bonding to paper cellulose.
Correct Answer: Partition between mobile solvent and aqueous layer on cellulose.
Q2. What does the Rf value represent in paper chromatography?
- The ratio of solute distance to solvent front distance travelled.
- The retention factor measured by peak area.
- The rate of flow of solvent in cm/min.
- The resolution factor between two spots.
Correct Answer: The ratio of solute distance to solvent front distance travelled.
Q3. Which development technique is best for separating volatile solvent systems and avoiding tailing due to evaporation?
- Ascending paper chromatography.
- Descending paper chromatography.
- Radial paper chromatography.
- Two-dimensional paper chromatography.
Correct Answer: Descending paper chromatography.
Q4. In partition-type paper chromatography, increasing the polarity of the mobile phase generally causes:
- Non-polar solutes to move further up the paper.
- Polar solutes to move further up the paper.
- No change in Rf values for any solute.
- Complete adsorption of all solutes to paper.
Correct Answer: Polar solutes to move further up the paper.
Q5. Which factor does NOT significantly affect Rf values in paper chromatography?
- Type and composition of solvent (mobile phase).
- Temperature during development.
- pH of the sample solvent.
- Color of the laboratory walls.
Correct Answer: Color of the laboratory walls.
Q6. How is the solvent front marked when the chromatogram is removed from the chamber?
- By estimating its position visually and drawing a line.
- By immediately drawing a pencil line at the solvent front.
- By measuring the distance from baseline to top of paper only later.
- By pressing the paper to transfer solvent to a glass slide.
Correct Answer: By immediately drawing a pencil line at the solvent front.
Q7. Which detection method is commonly used for non-UV active drugs on paper chromatograms?
- UV absorption at 254 nm.
- Chemical staining followed by visualization.
- Direct mass spectrometric imaging on paper.
- Refractive index detection of the paper.
Correct Answer: Chemical staining followed by visualization.
Q8. What is the main advantage of radial (circular) paper chromatography?
- Requires larger sample volumes than other methods.
- Allows faster separation with symmetric diffusion paths.
- Eliminates need for solvent selection.
- Prevents any band broadening entirely.
Correct Answer: Allows faster separation with symmetric diffusion paths.
Q9. Which statement about sample application in paper chromatography is true?
- Applying a very concentrated spot always improves resolution.
- Small, concentrated application minimizes band diffusion and improves resolution.
- Large volume application reduces tailing and improves sharpness.
- Applying sample in multiple layers with solvent evaporation can focus the spot.
Correct Answer: Applying sample in multiple layers with solvent evaporation can focus the spot.
Q10. In paper chromatography, what effect does chamber saturation with solvent vapor have?
- Increases reproducibility and minimizes solvent evaporation at the paper surface.
- Always decreases Rf values for all solutes.
- Prevents any movement of analytes on paper.
- Causes immediate precipitation of the sample.
Correct Answer: Increases reproducibility and minimizes solvent evaporation at the paper surface.
Q11. Which parameter is crucial when selecting a solvent system for resolving similar polar drug molecules?
- Relative polarity and elution strength of solvent mixture.
- Color of solvents used.
- Price of solvent only.
- Solvent boiling point exclusively.
Correct Answer: Relative polarity and elution strength of solvent mixture.
Q12. Why is pencil recommended to mark origin and solvent front on chromatograms?
- Pencil marks dissolve in solvents and indicate movement.
- Ink may run or interfere with the chromatogram, while pencil is inert.
- Pencil generates a UV-active line to aid detection.
- Pencil contains dyes that assist partitioning.
Correct Answer: Ink may run or interfere with the chromatogram, while pencil is inert.
Q13. Which is a limitation of paper chromatography in modern pharmaceutical analysis?
- Inability to separate any polar compounds.
- Lower resolution and reproducibility compared with HPLC.
- It is faster and more sensitive than HPLC in all cases.
- Complete automation is impossible.
Correct Answer: Lower resolution and reproducibility compared with HPLC.
Q14. How is quantitative estimation performed from paper chromatograms?
- Only by measuring Rf values.
- By densitometry or spot elution followed by spectrophotometric assay.
- By visually comparing colors only.
- By weighing the paper before and after chromatography.
Correct Answer: By densitometry or spot elution followed by spectrophotometric assay.
Q15. What causes tailing of spots in paper chromatography?
- Perfect sample application and optimal solvent.
- Strong interaction of analyte with stationary phase or overloading.
- Using highly porous silica paper only.
- Excessive chamber saturation always removes tailing.
Correct Answer: Strong interaction of analyte with stationary phase or overloading.
Q16. Which modification converts paper chromatography into a better reproducible partition system?
- Using untreated blotting paper only.
- Treating paper with a thin layer of liquid stationary phase (impregnation).
- Heating paper to high temperatures before use.
- Applying electrical field across the paper.
Correct Answer: Treating paper with a thin layer of liquid stationary phase (impregnation).
Q17. In two-dimensional paper chromatography, why is the plate rotated 90° for the second run?
- To use the same solvent twice for faster results.
- To separate spots that co-migrated in the first dimension using a different solvent.
- To make the paper look symmetrical.
- To evaporate residual solvent more quickly.
Correct Answer: To separate spots that co-migrated in the first dimension using a different solvent.
Q18. What is the role of pH in paper chromatographic separation of ionizable drugs?
- pH has no effect on ionizable drugs.
- It controls degree of ionization, altering partition behavior and Rf values.
- It only affects paper texture, not solute migration.
- It changes the cellulose structure irreversibly.
Correct Answer: It controls degree of ionization, altering partition behavior and Rf values.
Q19. Which practice improves resolution between two closely migrating spots?
- Applying a very large, concentrated sample spot.
- Optimizing solvent polarity, using narrower application and longer run distance.
- Reducing development distance to a minimum.
- Increasing paper thickness dramatically.
Correct Answer: Optimizing solvent polarity, using narrower application and longer run distance.
Q20. Preparative paper chromatography is used to:
- Quantify trace impurities at ppb levels only.
- Isolate and collect milligram to gram quantities of components separated on paper.
- Measure pH of drug solutions directly on paper.
- Replace all HPLC preparative separations.
Correct Answer: Isolate and collect milligram to gram quantities of components separated on paper.
Q21. Why is filter paper often preferred over ordinary paper for chromatography?
- Filter paper has a more uniform thickness and pore structure, improving reproducibility.
- Ordinary paper absorbs solvents faster so it is superior.
- Filter paper always contains silica which aids separation.
- Filter paper is colored to help visualization.
Correct Answer: Filter paper has a more uniform thickness and pore structure, improving reproducibility.
Q22. Which of the following indicates a partition mechanism rather than adsorption on paper?
- Dependence of migration on solvent polarity and relative water binding to cellulose.
- Irreversible binding of spot to cellulose fibers.
- Complete retention of all solutes at origin regardless of solvent.
- Spot migration always towards origin.
Correct Answer: Dependence of migration on solvent polarity and relative water binding to cellulose.
Q23. How can one improve sensitivity for detecting a faint spot on paper chromatography?
- Use a stronger stain or concentrate the eluted spot for spectrophotometric detection.
- Always use visible light only; UV is never helpful.
- Wash the paper with water to make spots clearer.
- Expose the paper to direct sunlight to intensify color.
Correct Answer: Use a stronger stain or concentrate the eluted spot for spectrophotometric detection.
Q24. Which solvent mixture strategy helps resolve closely related polar solutes?
- Start with a highly non-polar solvent and never change composition.
- Use binary or ternary mixtures and adjust polarity incrementally to achieve partial separation.
- Choose solvents with identical polarities only.
- Always use pure water as mobile phase.
Correct Answer: Use binary or ternary mixtures and adjust polarity incrementally to achieve partial separation.
Q25. What is the effect of temperature on paper chromatography separations?
- Increasing temperature always stops migration.
- Temperature can alter solvent viscosity and solute partition, changing Rf values and resolution.
- Temperature has no effect on partition equilibria.
- Lower temperatures convert partition to adsorption mechanisms.
Correct Answer: Temperature can alter solvent viscosity and solute partition, changing Rf values and resolution.
Q26. For ion-pair chromatography on paper, what is often added to the mobile phase?
- Non-ionic detergent only.
- An ion-pair reagent (counter ion) to form neutral complexes with ionic analytes.
- Strong acid to destroy paper cellulose.
- High concentrations of salt to saturate paper permanently.
Correct Answer: An ion-pair reagent (counter ion) to form neutral complexes with ionic analytes.
Q27. Which practice helps prevent streaking of polar compounds on paper?
- Applying sample as large wet spot to dilute concentration.
- Using diluted sample with repeated small applications and thorough drying between additions.
- Using extremely volatile solvents for application only.
- Heating the paper before application to high temperature.
Correct Answer: Using diluted sample with repeated small applications and thorough drying between additions.
Q28. How does paper chromatography compare with thin-layer chromatography (TLC) in pharmaceutical labs?
- Paper chromatography generally offers higher resolution than TLC in all cases.
- TLC is usually faster, more reproducible and more widely used for routine pharmaceutical separations.
- Paper chromatography can separate inorganic ions better than TLC always.
- Both techniques are identical in stationary phase chemistry.
Correct Answer: TLC is usually faster, more reproducible and more widely used for routine pharmaceutical separations.
Q29. Which troubleshooting step is recommended if solvent front is not moving uniformly along the paper?
- Ensure the paper is straight, chamber is level and solvent volume and saturation are correct.
- Increase sample load to force solvent flow.
- Remove solvent from chamber and apply heat directly to paper.
- Cut the paper into pieces to speed movement.
Correct Answer: Ensure the paper is straight, chamber is level and solvent volume and saturation are correct.
Q30. Which pharmaceutical application is paper chromatography still useful for today?
- High-throughput quantitation of complex formulations replacing HPLC.
- Simple qualitative screening for drug identity, impurity spot checks and educational demonstrations.
- Sterility testing for injectable products exclusively.
- Measuring tablet hardness directly on paper.
Correct Answer: Simple qualitative screening for drug identity, impurity spot checks and educational demonstrations.
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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