3.4. Sephadex LH-20 Column Chromatography

Low- and a high-molecular-weight (LMW and HMW) fractions from the MCE and ACE were collected using a Sephadex LH-20 chromatographic column [52]. A crude extract (200 mg) was dissolved in 95% (v/v) ethanol and loaded onto the column. Flow through the column was initiated by gravity using 95% (v/v) ethanol to elute the LMW fraction. The mobile phase was then changed over to 50% (v/v) aqueous acetone to recover the HMW fraction. Solvents were evaporated from collected fractions using the Rotavapor at 45 °C. The remaining aqueous residue was removed by lyophilization, and the product then stored in amber-glass vials at 4 °C. The prepared methanolic LMW and HMW fractions as well as acetonic LMW and HMW fractions are abbreviated as MLF, MHF, ALF, and AHF, respectively.