In situ assay of the reducing sugars in hydrophilic natural deep eutectic solvents by a modified DNS method

The DNS assay has been already a universally method for the in situ determination of reducing sugars (RS) in aqueous phase solvents. Today, "greener biorefineries" require in situ measurement of reducing sugars in nonaqueous solvents such as natural deep eutectic solvents (NADESs). Therefore, that DNS assay cannot be performed successfully in such eutectic phase solvent becomes a puzzle that must be solved. In this work, an improved DNS method, which we call the DNS" method, was established and implemented successfully in hydrophilic eutectic phase solvent systems (hiNADES). The tickets to the success of DNS" method are the inserted two essential steps, water-adding disintegration and pH alkalization. Then we tested two excellent RS solvents (ChCl_Gly and ChCl_LA) from six synthetic ChCl-based hiNADESs. The optimal disintegration conditions for assaying RS in ChCl_LA/NaOH (ChCl_Gly/H2O) were volume ratio 3: 1 (3: 1), molar concentration 2.0 mol/L (0 mol/L). Thereafter, we drew the standard curves of seven RSs in ChCl_Gly and ChCl_LA, which are glucose, xylose, fructose, mannose, fucose, rhamnose, and maltose. We determined that HMF interferes with DNS assay in hiNADES. The principles and standard procedures of DNS" method were also investigated. Finally, the enzymatic saccharification of microcrystalline cellulose (MCC) by two commercial Trichoderma reesei cellulases (Celluclast & REG; 1.5 L and Cellic & REG; CTec2) in neat hiNADES was tracked using the DNS" method.