thick. Commercially produced plates are available in several sizes between 5 cm
and 20 cm square and may incorporate an insoluble fluorescent reagent to facili-
tate the detection of solute spots (vide infra). The most commonly used sorbents
are silicaand powdered cellulose, and the corresponding sorption mechanisms
are adsorption and partition, respectively. Thin layers can also be made of
chemically-modified silicas, ion-exchange resins, exclusion gels and cyclo-
dextrins that display chiral selectivity. Some of these sorbents are similar to the
bonded phases used in HPLCwhich are discussed in Topics D6 and D7. Some
TLC sorbents are listed in Table 1.Table 1. Stationary phases (sorbents) for thin-layer chromatography
Sorbents Chromatographic mechanism Typical applications
Silica gels Adsorption Amino acids, hydrocarbons, alkaloids,
vitamins
Hydrocarbon modified silicas Modified partition Nonpolar compounds
Cellulose powder Partition Amino acids, nucleotides, carbohydrates
Alumina Adsorption Hydrocarbons, alkaloids, food dyes,
lipids, metal ions
Kieselguhrs (diatomaceous earths) Partition Sugars, fatty acids
Ion-exchange celluloses Ion-exchange Nucleic acids, nucleotides, halide and
metal ions
Sephadex gels Exclusion Polymers, proteins, metal complexes
b-Cyclodextrins Stereo-adsorptive interactions Mixtures of enantiomers
Mobile phase The range of mobile phases used in TLCis extremely wide and they are often
selected empirically. Blends of two solvents are common because the solvent
strength, or eluting power, can be easily adjusted to optimize a separation by
altering solute distribution ratios. Some general guidelines in selecting and
optimizing the composition of a mobile phase are
● Solvents should be of the highest purity as TLCis a very sensitive analytical
technique;
● Mobile phase eluting power should be adjusted so that solute Rfvalues fall
between 0.2 and 0.8 so as to maximize resolution;
● For separations on silica gel and other polar adsorbents, the overall polarity
of the mobile phase determines solute migration rates and hence their Rf
values; small additions of a slightly polar solvent, such as diethyl ether, to a
nonpolar solvent, such as methylbenzene, will significantly increase Rf
values;
● Polar and ionic solutes are best separated using a blend of a polar organic
solvent, such as n-butanol, with water; the addition of small amounts of
ethanoic acid or ammonia to the water increases the solubilities of basic and
acidic solutes, respectively.A helpful guide to solvent strength in adsorption and partition-based separa-
tions is an eluotropic series, an example of which is given for HPLCin Table 1
of Topic D6. The solvents are listed in order of increasing solvent strength for
adsorption-based separations. The order for partition-based separations is
broadly similar.D3 – Thin-layer chromatography 133