In this work, the use of glucose as feedstock to produce the biofuel 5-ethoxymethylfurfural (EMF) is explored using a two-step system. The first step is optimized for the dehydration of glucose into 5-hydroxymethylfurfural (HMF) using water:methyl isobutyl ketone (MIBK) and water:MIBK:ethanol biphasic systems, with the addition of CaCl2 and Beta-zeolites as catalysts for glucose dehydration. After optimization, up to a 61 % HMF yield is obtained after 30 min at 180 ◦C. Most of the HMF generated is extracted to the organic phase due to a high partition coefficient. This phase was used in a second step, by simple separation from the aqueous phase, with fresh catalyst. In the absence of water, the ethanol present with MIBK in the organic phase is enough to etherify the HMF to produce our desired biofuel. Etherification took place with a 67 % selectivity after 3 h at 140 ◦C. The yield of HMF could be improved this way respect to a one-step reaction, and a final EMF yield of 37 % EMF yield from glucose was achieved.