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Melamine (1,3,5-triazine-2,4,6-triamine or cyanuramide, C3H6N6) is a trimer of cyanamide, with a 1,3,5-triazine skeleton (Figure 3.5-1). The molecule contains 66% nitrogen by mass and, if mixed with resins, has fire retardant properties due to its release of nitrogen gas when burned or charred. The word melamine (from German) is a combination of the word melam (which is a distillation derivative of ammonium thiocyanate) and amine [1]. Melamine is also a metabolite of cyromazine, an insecticide in which the proton of an NH2-group is substituted by a cyclopropyl group.
High-performance thin-layer chromatography (HPTLC), as the modern form of TLC (thin-layer chromatography), is suitable for detecting pharmaceutically active compounds over a wide polarity range using the gradient multiple development (GMD) technique. Diode-array detection (DAD) in conjunction with HPTLC can simultaneously acquire ultraviolet‒visible (UV‒VIS) and fluorescence spectra directly from the plate. Visualization as a contour plot helps to identify separated zones. An orange peel extract is used as an example to show how GMD‒DAD‒HPTLC in seven different developments with seven different solvents can provide an overview of the entire sample. More than 50 compounds in the extract can be separated on a 6-cm HPTLC plate. Such separations take place in the biologically inert stationary phase of HPTLC, making it a suitable method for effect-directed analysis (EDA). HPTLC‒EDA can even be performed with living organism, as confirmed by the use of Aliivibrio fischeri bacteria to detect bioluminescence as a measure of toxicity. The combining of gradient multiple development planar chromatography with diode-array detection and effect-directed analysis (GMD‒DAD‒HPTLC‒EDA) in conjunction with specific staining methods and time-of-flight mass spectrometry (TOF‒MS) will be the method of choice to find new chemical structures from plant extracts that can serve as the basic structure for new pharmaceutically active compounds.