Three-Phase Partitioning for the Extraction and Purification of Polysaccharides from the Immunomodulatory Medicinal Mushroom Inonotus obliquus.

Polysaccharides from the immunomodulatory medicinal mushroom Inonotus obliquus (IOPS) were extracted and purified using three-phase partitioning (TPP), which is an efficient, fast, safe, and green purification technique. An optimal extraction procedure that gave a good 2.2% isolated yield was identified, using the following protocol: a solid-liquid ratio of 1 g to 12 mL; mass fraction of (NH₄)₂SO₄ 20% (w/v); 11 mL t-butanol; pH 8.0; temperature 30 °C; and extraction time 30 min. The purified IOPS was shown to be a proteoglycan of 40 kDa molecular weight comprising of d-galactose, d-glucose, d-xylose, and d-mannose in a molar ratio of 2.0:3.5:1.0:1.5. The purified IOPS displayed strong free-radical scavenging abilities, antioxidant activities, and immunological activity in vitro. IOPS' Trolox antioxidant equivalent capacity and ferric-reducing ability of plasma were 251.2 μmol Trolox/g sample and 1040.5 μmol Fe(2+)/g sample, respectively, with the activity of its immunomodulatory behavior shown to be gradient dependent.