TLC Determination of Aliphatic Polyamines on Calcium Sulfate Layers

Biogenic polyamines are sensitive markers for various diseases including cancer. Polyamines are difficult to analyze by chromatography due to their high polarity and water-solubility so that derivatization is an essential step for their chromatographic analysis. Earlier studies have shown the efficacy of calcium sulfate (CaSO₄) as a TLC coating material for the separation of polar compounds. The aim of this study was to explore the potential of CaSO₄ for the analysis of aliphatic polyamines without derivatization. The TLC of six polyamines (ornithine, citrulline, putrescine, cadaverine, spermidine and spermine) was carried out on CaSO₄ and silica gel plates using 11 different mobile phases. The results showed that CaSO₄ is superior to silica for the separation of underivatized polyamines. The development time of the CaSO₄ plates was also about one-third shorter as compared to silica. Methanol was the only solvent to produce differential R _F values for the polyamines studied. Ornithine (R _F , 0–2) and citrulline (R _F , 1–3) were separated from cadaverine (R _F , 0.93), spermine (R _F , 0.85) and spermidine (R _F , 0.85). For quantitative analysis, the polyamines were eluted from the coating material scratched from the plate and the absorbance of the supernatant was measured at 550 nm. The limits of detection (LOD) and quantification (LOQ) were found to be 0.75 and 1.88 μg, respectively. The procedure was applied to the quantitative separation of polyamines in spiked human urine samples (12.5–50 μg). This is probably the first study reporting a TLC method for the separation of underivatized polyamines.