Simultaneous determination of naphthalimide-labelled monosaccharides in P. cyrtonema Hua. polysaccharides utilizing the HPLC-UV technique

Abstract

4-Amidogen-1,8-naphthalimide (ANA), a novel pre-column derivatization reagent, has been successfully synthesized and utilized for the highly sensitive analysis of monosaccharides. ANA reacts with the reducing carbonyl groups of saccharides, facilitating monosaccharide detection. The resulting monosaccharide derivatives were meticulously investigated using High-Performance Liquid Chromatography (HPLC) coupled with ultraviolet (UV) spectroscopy. The derivatization process was conducted in an environmentally friendly solvent mixture of EtOH and AcOH at a constant temperature of 100 °C for a duration of 4 hours across all monosaccharides. After derivatization, the separated adducts were eluted through a ZORBAX SB-C₈ column within a mere 22 minutes at a flow rate of 0.8 mL min⁻¹, with detection occurring at a wavelength of 416 nm. By employing this derivatization technique, the comprehensive analysis of the monosaccharide composition in P. cyrtonema Hua. polysaccharides (PCPs) following strong acid hydrolysis was achieved. The monosaccharide profiles of PCPs predominantly consist of glucose (Glc), galactose (Gal), mannose (Man), xylose (Xyl), and galacturonic acid (GalA), with a molar ratio of Glc : Gal : Man : Xyl : GalA of 20.26 : 12.76 : 24.24 : 41.91 : 0.83. The LOD values for the six sugars ranged from 27.37 to 84.37 μg mL⁻¹. The validated HPLC method, characterized by its exceptional precision and accuracy, has successfully facilitated both rapid qualitative and quantitative determination of PCPs, thereby establishing a robust quality control protocol for P. cyrtonema Hua.