Ionic Liquid Pretreatment in Tandem with Recombinant Agarase Cocktail Saccharification of Gelidium amansii for D-Galactose and 3,6-Anhydro-L-Galactose Production

D-Galactose (D-Gal) and 3,6-Anhydro-L-galactose (3,6-L-AHG) are valuable sugars that can be derived from Gelidium amansii. Herein, ionic liquid (IL) pretreatment was used to deconstruct the cell wall of G. amansii without producing any side-products from sugar degradation. It is combined with enzymatic saccharification using recombinant agarases to effectively produce D-Gal and 3,6-L-AHG from IL-treated G. amansii. ILs were screened for the biomass pretreatment as well as the antisolvents for dissolved biomass retrieval from the IL. [Bmim]Ac is the most effective solvent (99% dissolution) and methanol as antisolvent (78% reconstituted G. amansii). Using simplex-centroid design, the optimal loading of three beta-agarases: Aga2, AgaA7, and Aga50D for the first hydrolysis was determined to be at equal fractions (0.33). Addition of alpha-neoagarobiose hydrolase AhgI finally produced maximum yields of 56.5% for D-Gal and 33.7% for 3,6-L-AHG. Given the industrial importance of D-Gal and the high market price of 3,6-L-AHG, results demonstrate the potential of combined IL pretreatment and enzymatic hydrolysis using recombinant agarases as a green process for sugar production from red macroalgae.