Ultrasound-assisted deep eutectic solvent extraction combined with gradient ethanol precipitation of polysaccharides from Paulownia fortunei flowers: Process optimization, physicochemical characterization, bioactivity evaluation.

The traditional methods for extraction and separation of polysaccharides usually have the shortcomings of long processing time and complex operation procedures. The objective of this study was to develop an efficient and integrated strategy for synergistic extraction and separation of polysaccharides from Paulownia fortunei flowers by ultrasound-assisted deep eutectic solvents extraction (UADES) combined with gradient ethanol precipitation (GEP). The extraction conditions were successively optimized using single-factor experiments and response surface methodology. Under the optimal conditions, the maximum extraction yield of P. fortunei flowers polysaccharides (PFP) reached 5.91 ± 0.29%, which was 3.05-fold higher than that of conventional hot water extraction (CHWE). Analysis of tissue morphology, and molecular dynamics simulations of DES and PFP both indicated that this extraction method was superior to CHWE. Meanwhile, PFP can be fractionated and purified through GEP. Three polysaccharide fractions, namely PFP-40, PFP-60, and PFP-80, were obtained at the final ethanol precipitation concentrations of 40%, 60%, and 80% (v/v), respectively. All fractions also showed differences in structural characterizations. In vitro activity assays demonstrated that all polysaccharide fractions exhibited antioxidant activity and tyrosinase inhibitory activity, and PFP-80 showed the strongest bioactivity. Further mechanistic studies indicated that the inhibition of tyrosinase by PFP-80 was reversible and mixed-type, their interaction occurred through a static quenching mechanism. This study introduced a strategy for DES recycling post-GEP for polysaccharide extraction, and the PFP showed potential for applications in the food and pharmaceutical industries.