Abstract The effect of a synergized two-step subcritical water extraction (SWE) and simultaneous multi-frequency ultrasound-assisted alcohol/salt aqueous two-phase extraction (MFu-AATPE) on the yield, physicochemical, structural characteristics, and antioxidant activities of Lentinus edodes polysaccharides (LEPs) was investigated. Crude LEP extracted under subcritical extraction (31.14%) was purified using dual and triple frequency ultrasound-assisted aqueous two-phase systems giving a yield of 94.57% and 97% respectively. Compared with the dual-frequency sonication (20/40 kHz), the triple frequency sonication (20/40/60 kHz) gave maximum yield and improved the desalination rate by 33.6%. Congo red analysis revealed triple helix structures. Gas Chromatography analysis showed the existence of mannose, glucose, arabinose, xylose, and galactose whilst spectral similarities were observed through Fourier Transform Infrared Spectroscopy. The average molecular weight (Mw) was 83.6170 Da, 2384 Da and 2387 Da for control, dual and triple frequency samples respectively. Ultrasound treated polysaccharides displayed stronger antioxidant activities against the 2.2-diphenyl;-1 picrylhydrazyl (DPPH), 2, 2ʹ-azino-bis 3-ethylbenzothiazoline-6 sulfonic acid (ABTS) and hydroxyl radicals, thus demonstrating their potency in reducing oxidation. However, the microstructure of the triple frequency treated sample was altered compared to the dual-frequency sample. Therefore, this study demonstrated the efficacy of SWE in crude LEPs extraction and illustrated MFu-AATPE as an efficient technique that selectively isolates polysaccharides and enhances extraction efficiencies.

中文翻译：

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蘑菇多糖的协同亚临界超声辅助水相两相萃取、纯化和表征

摘要 协同两步亚临界水萃取 (SWE) 和同时多频超声辅助醇/盐水溶液两相萃取 (MFu-AATPE) 对香菇的产量、理化、结构特征和抗氧化活性的影响香菇多糖 (LEP) 进行了研究。在亚临界萃取下萃取的粗 LEP (31.14%) 使用双频和三频超声辅助水性两相系统进行纯化，产率分别为 94.57% 和 97%。与双频超声（20/40 kHz）相比，三频超声（20/40/60 kHz）产量最大，脱盐率提高了33.6%。刚果红分析揭示了三螺旋结构。气相色谱分析表明存在甘露糖、葡萄糖、阿拉伯糖、木糖、和半乳糖，同时通过傅立叶变换红外光谱观察到光谱相似性。对照、双频和三频样品的平均分子量 (Mw) 分别为 83.6170 Da、2384 Da 和 2387 Da。超声处理的多糖对 2.2-diphenyl;-1picrylhydrazyl (DPPH)、2, 2ʹ-azino-bis 3-ethylbenzothiazoline-6 磺酸 (ABTS) 和羟基自由基显示出更强的抗氧化活性，从而证明了它们在减少氧化方面的效力。然而，与双频样品相比，三频处理样品的微观结构发生了变化。因此，本研究证明了 SWE 在粗 LEP 提取中的功效，并说明 MFu-AATPE 作为一种有效的技术，可以选择性地分离多糖并提高提取效率。