This study is a first attempt to present a report about a high-performance thin-layer chromatography (HPTLC)–densitometric method that has been developed and validated for the quantification of sugars, i.e., glucose, fructose, sucrose, maltose, and raffinose from the methanol extracts of roots of 7 Asparagus species ‒ A. adscendens, A. racemosus, A. retrofractus, A. officinalis, A. densiflorus, A. falcatus, and A. sprengeri. The separation and quantification of sugars were achieved using the mobile phase propanol‒ethyl acetate‒ water (6:3:1, v/v). Derivatization of the bands was done by using diphenylamine, aniline, and ortho-phosphoric acid reagents, and densitometric scanning was done at 600 nm in absorbance–reflectance mode. This method gave the best resolution of bands at RF = 0.55, 0.50, 0.31, 0.11, and 0.46, corresponding to fructose, glucose, maltose, raffinose, and sucrose, respectively. The calibration curves were linear in the range of 100–500 ng per spot. The limit of detection (LOD) and the limit of quantification (LOQ) ranged between 12.77–75.48 ng and 38.69–228.73 ng, respectively. The intraand inter-day precisions were found between 0.33–1.46 in terms of relative standard deviation (RSD). The average recoveries ranged between 97.84 –99.91%. The validated method was successfully used to detect and quantify sugars in the methanol root extracts of the 7 Asparagus species.
Validation of analytical procedure: methodology Q2 (R1), International Conference on Harmonization (ICH), International Federation of Pharmaceutical Manufacturers and Associations (IFPMA), Geneva, 1996.