High performance thin-layer chromatography Dušanka Milojković-Opsenica* and Filip Andrić Faculty of Chemistry, University of Belgrade, Studentski trg 12-16, 11000 Belgrade, Serbia Corresponding author: dusankam@chem.bg.ac.rs Abstract The current chapter emphasizes the use of high-performance thin- layer chromatography (HPTLC) as a green analytical separation al- ternative. The chapter provides basic insight into terms and histori- cal development of thin-layer chromatography towards becoming modern, automated, high resolution technique in the form of high- performance thin-layer chromatography, and their further advances in miniaturization of chromatographic beds in the form of ultra- performance thin-layer chromatography (UPTLC). Also, following the three R philosophy (reduce, replace, reuse), and twelve prin- ciples of green chemistry, the chapter provides the short review of green sample pretreatment techniques coupled with the HPTLC. In addition, the use of green separation modalities such as reversed- phase chromatography, chromatography of hydrophilic interaction and salting out thin-layer chromatography is emphasized. The use of thin-layer chromatography as a simple screening technique and a powerful sample pretreatment method is encouraged.