Recovery of steviol glycosides from industrial stevia by-product via crystallization and reversed-phase chromatography

Mother liquor sugar (MLS), as the by-product of stevia production, contained similar to 65% steviol glycosides (SGs). Recovery of the SGs from MLS was achieved by crystallization coupled with reversed-phase chromatography. Crystallization was done by dissolving MLS in methanol solution and SGs were crystallized due to the polarity difference from the medium. Composition of SGs crystals differed with various temperature, time, solid-to-liquid ratio and water content. SGs were 42.25% recovered with high purity of 96.89% under optimal conditions (solid-to-liquid ratio = 1:5 (w/v), T = 25 degrees C, t = 24 h) in absolute methanol. The liquid phase after crystallization was subsequently subjected to reversed-phase chromatography, whereby the impurities were firstly eluted with 35% (v/v) ethanol solution and the purified SGs were then desorbed by absolute ethanol, finally recovering 95.20% of SGs in the purity of 98.08%. The total SGs recovery of the whole procedure was 97.23%. The two-step purification was easy-to-operate and feasible to scale-up for industrial application.