A simple, selective, precise, and reproducible high-performance thin-layer chromatography (HPTLC) method for the analysis of vicine in herbal extracts and formulations containing Momordica charantia was developed. The study was performed on aluminum-based pre-coated TLC plates (silica gel 60F254). The chromatograms of samples were developed in twin trough glass chamber pre-saturated with mobile phase comprised of ethyl acetate-methanol-water-formic acid (7.5:3:1:0.1, v/v/v/v) at room temperature (25 ± 2°C). The densitometric analysis was carried out in absorbance mode at 278 nm. The optimized mobile phase showed a compact spot of vicine (RF = 0.31 ± 0.02) from the other constituents present in the samples. The linear regression analysis data for the calibration plots showed good linearity (r2 = 0.9990) with respect to peak area in the range of 200–1400 ng spot -1. The method was validated as per International Conference on Harmonization (ICH) guidelines. The limits of detection and quantification (50 and 150 ng spot−1, respectively) were also established. The proposed method has shown the excellent recovery (98.01–99.33%), which supports the suitability of the method for the analysis. Statistical analysis of the obtained data showed the selectivity of the proposed method for vicine estimation in the herbal samples.
Zmodyfikowano metodę HPLC oznaczania glikopiranozydów (wicyny i konwicyny) poprzez zastosowanie dostępnej w handlu urydyny jako standardu wewnętrznego. Wyznaczone doświadczalnie współczynniki korekcyjne wynosiły dla wicyny i konwicyny odpowiednio 1,12 i 0,79. Prześledzono zmiany zawartości wicyny i konwicyny w czasie dojrzewania nasion bobiku stwierdzając ponad 90% redukcję ich zawartości w nasionach bobiku w czasie potrzebnym do osiągnięcia dojrzałości przemysłowej.
EN
Uridine (C9H12N2O6) was applied as an internal standard for the determination of glucopiranosides in faba bean seeds by the HPLC method. The relative UV response factors of vicine or convicine to uridine were determined (1.12 and 0.79, respectively). The changes in the content of vicine and convicine during ripening of faba bean seeds were followed. It was observed that with the increase in the seed dry matter content from 0.18 kg/kg to 0.78 kg/kg, the dry matter based content of vicine decreased from 26.225 g/kg to 0.243 g/kg, and that of convicine from 9.051 g/kg to 0.094 g/kg.