Paper chromatography is a classical, cost-effective method for the separation of sugars by exploiting differences in adsorption to the stationary cellulose paper and solubility in the mobile solvent. B. Pharm students must grasp core concepts: stationary phase (Whatman/cellulose), mobile phase selection and polarity, capillary action-driven development, sample spotting, solvent front, Rf values, and visualization reagents (Molisch’s, anthrone). Understanding factors affecting resolution—solvent composition, temperature, sample load, and two‑dimensional chromatography—helps in qualitative identification and semi‑quantitative analysis of mono- and disaccharides in pharmaceutical samples. Practical mastery supports quality control, excipient profiling, and formulation troubleshooting. Now let’s test your knowledge with 30 MCQs on this topic.
Q1. What is the stationary phase in conventional paper chromatography for sugars?
- Silica gel coated on paper
- Cellulose fibers of Whatman paper
- Alumina powder impregnated on paper
- Cellulose acetate membrane
Correct Answer: Cellulose fibers of Whatman paper
Q2. Which property of the solvent primarily determines the separation of polar sugars in paper chromatography?
- Density of the solvent
- Polarity of the solvent
- Viscosity of the solvent
- Boiling point of the solvent
Correct Answer: Polarity of the solvent
Q3. How is the Rf value calculated in paper chromatography?
- Distance moved by solvent front ÷ distance moved by solute spot
- Distance moved by solute spot ÷ distance from origin to solvent front
- Distance moved by solute spot ÷ distance moved by solvent front
- Distance from origin to solvent front ÷ distance moved by solute spot
Correct Answer: Distance moved by solute spot ÷ distance moved by solvent front
Q4. Which reagent gives a positive purple/violet reaction with carbohydrates on paper chromatograms?
- Ninhydrin
- Molisch’s reagent (α-naphthol + H2SO4)
- Dragendorff’s reagent
- Iodine vapor
Correct Answer: Molisch’s reagent (α-naphthol + H2SO4)
Q5. Two-dimensional paper chromatography is mainly used to:
- Reduce development time by half
- Increase sample loading capacity
- Separate complex mixtures with overlapping Rf values
- Eliminate need for visualization reagents
Correct Answer: Separate complex mixtures with overlapping Rf values
Q6. What is the usual purpose of drawing the origin line with a pencil before spotting samples?
- Pencil contains chemicals that enhance spots
- To avoid solvent interactions that would occur with ink
- Pencil line dissolves and migrates with sample
- Pencil prevents capillary action
Correct Answer: To avoid solvent interactions that would occur with ink
Q7. Which factor will NOT typically affect Rf values in paper chromatography?
- Ambient temperature during development
- Concentration of sample spotted
- Type of pencil used for origin line
- Composition of mobile phase
Correct Answer: Type of pencil used for origin line
Q8. Which solvent system is commonly used for separating monosaccharides on paper chromatography?
- Hexane : ethyl acetate
- Butanol : acetic acid : water (4:1:5)
- Methanol : chloroform
- Acetone : benzene
Correct Answer: Butanol : acetic acid : water (4:1:5)
Q9. Why is the solvent front marked immediately after development?
- To estimate sample concentration
- To prevent solvent evaporation from altering Rf measurement
- To enhance spot color intensity
- To fix the chromatogram permanently
Correct Answer: To prevent solvent evaporation from altering Rf measurement
Q10. Which visualization method is preferred for quantifying hexose sugars on paper chromatography?
- Molisch’s qualitative spray
- Anthrone reagent for colorimetric estimation
- Ninhydrin spraying
- UV light at 254 nm
Correct Answer: Anthrone reagent for colorimetric estimation
Q11. What does a low Rf value indicate about a sugar’s affinity in paper chromatography?
- High affinity for the mobile phase
- High affinity for the stationary phase
- The spot is unstable and degraded
- The solvent was too polar
Correct Answer: High affinity for the stationary phase
Q12. In a polar mobile phase, which sugar type tends to move farther up the paper?
- Highly polar sugars with strong hydrogen bonding
- Less polar sugars with weaker adsorption to paper
- Only disaccharides move farther
- Only reducing sugars move farther
Correct Answer: Less polar sugars with weaker adsorption to paper
Q13. Which paper grade is typically recommended for reproducible paper chromatography?
- Glossy copier paper
- Whatman filter paper no. 1 or equivalent
- Newspaper
- Cellophane sheet
Correct Answer: Whatman filter paper no. 1 or equivalent
Q14. Why is paper chromatography considered chromatographic despite lack of coated stationary phase?
- Because it uses liquid mobile phase and differential partitioning between phases
- Because paper is heated to create a stationary layer
- Because compounds are covalently bound to paper
- Because it requires high pressure for development
Correct Answer: Because it uses liquid mobile phase and differential partitioning between phases
Q15. Which of the following improves resolution between two close sugar spots?
- Increase spot size and concentration
- Use a weaker solvent that increases retention
- Use a paper with larger pore size
- Develop at a higher temperature without changing solvent
Correct Answer: Use a weaker solvent that increases retention
Q16. What is the main advantage of paper chromatography for sugar analysis in teaching labs?
- High sensitivity for trace analysis
- Low cost and simple setup with clear visual separation
- Requires no reagents for visualization
- Automated data output for quality control
Correct Answer: Low cost and simple setup with clear visual separation
Q17. Which property of sugars influences their migration order in paper chromatography?
- Molecular color only
- Molecular weight, polarity, and hydrogen-bonding ability
- Magnetic susceptibility
- Electrical conductivity
Correct Answer: Molecular weight, polarity, and hydrogen-bonding ability
Q18. What corrective step should be taken if sample spots tail during development?
- Increase sample volume applied
- Decrease sample concentration and use smaller, well-dried spots
- Change paper to cotton cloth
- Use hotter solvent to speed development
Correct Answer: Decrease sample concentration and use smaller, well-dried spots
Q19. How does pre-saturating the chromatography chamber with solvent vapor affect results?
- Causes uneven solvent front
- Reduces evaporation and gives more reproducible development
- Prevents capillary action altogether
- Removes all dissolved impurities from solvent
Correct Answer: Reduces evaporation and gives more reproducible development
Q20. Which detection reagent specifically differentiates pentoses from hexoses on paper chromatography?
- Molisch’s reagent
- Orcinol-sulfuric acid reagent
- Ninhydrin
- Biuret reagent
Correct Answer: Orcinol-sulfuric acid reagent
Q21. In paper chromatography, what is the origin?
- The top edge of the paper where solvent exits
- The line where sample spots are applied
- The point where solvent evaporates first
- The position of the detector
Correct Answer: The line where sample spots are applied
Q22. Which error would most likely produce artificially high Rf values?
- Allowing the solvent front to travel only a short distance
- Running the experiment at a lower temperature
- Touching and disturbing the paper during development
- Marking the solvent front incorrectly closer to origin
Correct Answer: Touching and disturbing the paper during development
Q23. What is the role of acetic acid in butanol:acetic acid:water solvent systems?
- Acts as an oxidizing agent
- Modifies ionization and hydrogen-bonding to improve separation of polar compounds
- Precipitates sugars onto paper
- Increases the viscosity to slow development
Correct Answer: Modifies ionization and hydrogen-bonding to improve separation of polar compounds
Q24. Which practice ensures more accurate Rf comparisons between runs?
- Using different paper types for each run
- Keeping solvent composition, paper grade, and temperature constant
- Varying sample spot size intentionally
- Changing chamber saturation each time
Correct Answer: Keeping solvent composition, paper grade, and temperature constant
Q25. For semiquantitative estimation of a sugar by paper chromatography, which approach is commonly used?
- Compare spot intensity to standard spots of known concentration using densitometry or colorimetric response
- Measure spot migration time only
- Estimate concentration by spot width alone
- Use mass spectrometry directly on the paper without standards
Correct Answer: Compare spot intensity to standard spots of known concentration using densitometry or colorimetric response
Q26. Which precaution should be taken when using acid reagents (e.g., anthrone) to visualize sugars?
- No precautions needed; they are inert
- Handle with care under fume hood; they are corrosive and can char paper
- Store reagents in direct sunlight to activate
- Mix with alkaline dye to neutralize before use
Correct Answer: Handle with care under fume hood; they are corrosive and can char paper
Q27. How does increasing the proportion of water in a butanol:acetic acid:water solvent typically affect sugar migration?
- Decrease polarity of solvent and increase migration
- Increase polarity and decrease migration of polar sugars
- Convert paper chromatography into gas chromatography
- Cause immediate precipitation of sugars at origin
Correct Answer: Increase polarity and decrease migration of polar sugars
Q28. Which statement correctly contrasts paper chromatography with thin-layer chromatography (TLC) for sugars?
- Paper chromatography uses silica; TLC uses cellulose
- TLC generally provides faster development and higher resolution due to coated stationary phases, while paper is simpler and cheaper
- TLC cannot separate sugars at all
- Paper chromatography requires UV lamps while TLC never does
Correct Answer: TLC generally provides faster development and higher resolution due to coated stationary phases, while paper is simpler and cheaper
Q29. What is the consequence of overloading a spot with too concentrated sugar solution?
- Sharper, more defined spots
- Spot diffusion, tailing, and poor resolution
- Completely prevents solvent movement
- Increases Rf uniformly for all compounds
Correct Answer: Spot diffusion, tailing, and poor resolution
Q30. Which experimental control helps confirm the identity of an unknown sugar on a chromatogram?
- Run a reference standard of the suspected sugar alongside the unknown under identical conditions
- Use a different solvent for the standard and unknown
- Spot the unknown far from the origin only
- Visualize with a reagent that reacts with proteins
Correct Answer: Run a reference standard of the suspected sugar alongside the unknown under identical conditions
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