Method development and solvent optimization for the separation of pharmaceutically and biologically important compounds by high-performance planar chromatography and automated multiple development

The multiple development technique in thin-layer chromatography (TLC) is a modern concept and versatile strategy which enables high spot capacities to be attained and is suitable for the analysis of very complex mixtures. In this work automated multiple development and high performance thin-layer chromatography (AMD-HPTLC) was used for the separation of pharmaceutically and biologically important compounds.; Automated multiple development is a microprocessor controlled instrumental technique for the stepwise solvent gradient development in thin-layer chromatography. The stepwise gradient is generated by changing the composition of the mobile phase for some or each successive cycle. The gradient usually commences with the strongest solvent and proceeds in successive cycles to solvents of lower strength. The principal function of the first solvent is band focusing, since it is applied for only a short development distance, and all sample components should have weak retention in this solvent. In subsequent development cycles the development distance is longer and the mobile phase weaker. Eventually the strength of the mobile phase will decline to the point that a particular sample component will no longer migrate and will occupy its final position in the chromatogram. The position and width of this zone is then hardly influenced by the further development cycles required to separate those substances which interact less strongly with the stationary phase. In this way the zones occupy characteristic positions within the chromatogram and have widths which are, to a larger extent, approximately equivalent throughout. Optimization of AMD involves two types of variables. The first includes the process variables which control the operation of the AMD device and includes all the time-based functions which must be set to execute a program. The second relates to the optimization of sample resolution and concerns the selection of both the appropriate stationary phase and the mobile phase constituents from which the most effective solvent gradient may be generated.; Solvent selection for suitable mobile phase gradients was made using the PRISMA solvent optimization model. Also, scanning densitometer was exclusively used for all detection purposes.; The results of this dissertation research show the effective application of multiple development thin-layer chromatography to the separation of poly(ethylene glycols), estrogens, protein PTH-amino acid derivatives, polar aromatic flavor compounds in vanilla extracts, food, and beverages, and major carboxylic acids present in wine. It also demonstrates that automated multiple development offers convenience, a high zone capacity, and high sample throughput.