The aim of the work was the chromatographic separation of salicylic acid and their derivatives, i.e., acetylsalicylic acid, salicylanilide, salicylaldehyde, salicylamide, methyl salicylate, phenyl salicylate, 2,5-dihydroxybenzoic acid, salicylhydroxamic acid, 3,5-dinitrosalicylic acid, 3-aminosalicylic acid, 4-aminosalicylic acid, and 5-aminosalicylic acid by use of adsorption thin-layer chromatography (normal-phase thin-layer chromatography [NPTLC]) and partition thin-layer chromatography (reversed-phase thin-layer chromatography/high-performance thin-layer chromatography [RP-TLC/HPTLC]). Three qualitatively and quantitatively different mobile phases were used for the separation of salicylic acid and its derivatives. Cluster analysis (single linkage method, Euclidean distance) allowed the evaluation of the suitability of the chromatographic conditions used to separate the pairs of tested compounds. The cluster analysis data indicate that the composition of the mobile phase is fundamental in the process of separation of the analyzed compounds by use of NP-TLC. The best separation of the studied substances was observed in the case of mobile phase n-hexane—diethyl ether—acetic acid (80%) in different volume ratios. The similarity analysis of the results obtained by use of RP-TLC/HPTLC revealed that the type of chromatographic plates influences significantly the quality of separation of the tested compounds. The best conditions for the separation by RP-TLC were obtained on silica gel RP-18 F254 plates. The present study indicates that the cluster analysis represents a simple-to-use and powerful chemometric tool in the prediction of TLC separation of medically important salicylic acid derivatives under various chromatographic conditions. It can be helpful in the quality control of multicomponent synthetic preparations containing these compounds or in the chemical standardization of plant products consisting of salicylic acid and related compounds.
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