Influence of sample preparation methods on the quantitation of selected tropane alkaloids from herb of Datura innoxia mill. by HPTLC

The aim of our paper was the examination of various procedures of extraction and purification methods on the yield of atropine and scopolamine in Datura innoxia extracts determined by high-performance thin-layer chromatography (HPTLC). In our experiments, the following extraction methods were used: percolation, ultrasound-assisted extraction (USAE) at various temperatures, and microwave-assisted extraction (MASE). The highest extraction yields of atropine and scopolamine were obtained by ultrasound-assisted extraction method and aqueous acetic acid or aqueous tartaric acid used as extractants. Crude extracts of D. innoxia herb were purified by use of various methods: solid-phase extraction-strong cation exchanger (SPE-SCX), SPE-C18, and liquid-liquid extraction (LLE). The best results of purification with satisfactory %RSD values were obtained by liquid-liquid extraction by use of dichloromethane from aqueous crude alkaloid extract alkalized to pH 12. HPTLC was performed on pre-coated plates Kieselgel 60 F254 in horizontal DS chambers by use of mobile phase: methanol-acetone-aqueous ammonia (5:4.5:0.5). The alkaloids’ content was determined in extracts by use of calibration curve method for atropine and scopolamine. Calibration curves were linear in range of 3.0–30 μg for scopolamine (R = 0.9980) and 2.6–26 μg for atropine (R = 0.9923). Limits of detection (LOD) and limits of quantification (LOQ) were 1.12 μg and 3.42 μg per spot for scopolamine and 0.89 μg and 2.82 μg per spot for atropine.