Ultrasound-Assisted green extraction and resin purification of Hypaphorine from Nanhaia speciosa using deep eutectic solvents.

Hypaphorine (HYP), a tryptophan-derived alkaloid with diverse pharmacological activities, was efficiently extracted from Nanhaia speciosa using a green and sustainable approach integrating ultrasound-assisted extraction (UAE), deep eutectic solvents (DESs), and macroporous resin (MAR) purification. The ultrasound-assisted DES extraction process was systematically optimized, with the best conditions identified as a lactic acid-fructose (1:1, mol/mol) DES containing 40 % water, an extraction temperature of 80 °C, a duration of 40  min, and a solid-to-liquid ratio of 1:20  g/mL. Under these conditions, the HYP yield reached 3.15  mg/g, representing improvements of 105.19 % and 70.62 % compared with conventional water and ethanol extraction, respectively, and 52.17 % higher than microwave-assisted extraction (MAE). To enhance purity, the crude extract was further purified using DM132 macroporous resin under optimized conditions (50 % ethanol eluent, 2.5 BV/h flow rate, column height-to-diameter ratio of 21:1, and elution volume of 4 BV), achieving a recovery rate of 89.53 % and a final purity of 79.57 %. DES recycling retained 94.21 % of the extraction efficiency after one reuse cycle. SEM analysis revealed significant disruption of the plant cell structure post-ultrasound-DES treatment. FTIR and DFT studies demonstrated strong hydrogen bonding and favorable binding energies between DES components and HYP, elucidating the extraction mechanism. This UAE-DES-MAR system not only enhanced the extraction and purification efficiency of HYP but also presented a green, recyclable, and scalable approach, offering promising applications for the sustainable industrial production of high-value alkaloids.