A chemist runs a silica TLC plate with hexane as the eluent and observes that a polar compound barely moves from the origin (Rf ≈ 0.05). What change to the eluent would best increase the compound's Rf to the target range of 0.3–0.5?
AAdd a small amount of a more polar solvent (e.g., ethyl acetate) to the hexane
BUse a less polar stationary phase (alumina instead of silica)
CIncrease the volume of hexane in the chamber to saturate the atmosphere
DRe-spot the sample with a higher concentration
In normal-phase TLC (polar silica stationary phase), polar compounds stick tightly to the silica and need a polar mobile phase to compete with them for the silica surface. Adding a more polar solvent like ethyl acetate to hexane increases the mobile phase's ability to pull the polar compound off the silica, raising its Rf. Chamber saturation (option C) helps reproducibility but doesn't change the relative polarity. Sample concentration (option D) affects spot darkness but not Rf. Option B would reduce stationary-phase polarity which could help, but changing the solvent is the standard first approach.
Question 2 Multiple Choice
Two unknown compounds are co-spotted on a silica TLC plate and run in ethyl acetate/hexane. Both spots comigrate at Rf = 0.42. What is the most appropriate conclusion?
AThe compounds are identical, since they have the same Rf under identical conditions
BThe compounds are identical only if they also show the same color under UV light
CThe compounds may be identical or different — same Rf in one system is insufficient to confirm identity; multiple eluent systems are needed
DThe compounds are definitely different, because two distinct substances cannot have the same Rf
Same Rf in a single solvent system does not confirm identity — different compounds can have the same Rf by coincidence if their overall polarity happens to be similar. Confirmation requires co-spotting in at least 2–3 different solvent systems with different polarity characteristics. If the spots comigrate in all systems, identity becomes much more likely (though NMR or mass spectrometry provides definitive confirmation). This is a classic TLC pitfall: comigration ≠ same compound.
Question 3 True / False
Increasing the polarity of the mobile phase in a normal-phase (silica) TLC experiment will increase the Rf values of the compounds being separated.
TTrue
FFalse
Answer: True
In normal-phase TLC, compounds compete with the mobile phase for adsorption sites on the polar silica stationary phase. A more polar mobile phase competes more effectively with compounds for silica binding, pulling them along with the solvent front and increasing their Rf values. This is why switching from hexane (nonpolar) to ethyl acetate (polar) causes spots to move farther up the plate, and why a very polar solvent like methanol pushes everything near the solvent front (Rf → 1).
Question 4 True / False
An Rf value measured in one experiment can be reliably compared to Rf values reported in the literature from a different laboratory to confirm compound identity.
TTrue
FFalse
Answer: False
Rf values are not absolute constants — they depend on adsorbent activity (how hydrated the silica is), mobile phase saturation of the chamber, temperature, plate batch, and solvent purity. Even small differences in these conditions between labs will shift Rf values. The correct approach is to always co-spot an authentic reference standard on the same plate in the same run, so any variation in conditions affects both compounds equally. Comparing raw Rf numbers across different experiments or laboratories is unreliable.
Question 5 Short Answer
Why must you co-spot an unknown compound with an authentic standard on the same TLC plate in the same run rather than comparing Rf values from separate experiments?
Think about your answer, then reveal below.
Model answer: Rf values shift with adsorbent activity, chamber saturation, temperature, and solvent composition. Co-spotting ensures both compounds experience identical conditions, making Rf comparison meaningful. Comparing values from separate runs introduces experimental variation that can make different compounds appear identical or the same compound appear different.
The key misconception to avoid is treating Rf as a physical constant like a melting point. It is a ratio measured under experimental conditions that are difficult to exactly replicate. By running the unknown and standard side by side on the same plate, you eliminate all sources of inter-run variability — any differences in spot position are then meaningful rather than artifacts of condition changes.