Short polyols, such as ethylene glycol (EG), are a popular target of catalytic hydrogenolysis of saccharides. However, studies on the use of untreated or pretreated lignocellulosic biomass as feedstock for polyol production are scarce. In this work, we have studied the impact of lignin on the catalytic hydrogenolysis of different biomass samples, targeting ethylene glycol. We first developed a hydrogenolysis protocol that is sensitive to lignin and feedstock impurities, such as ash and extractives. A matrix of biomass feedstocks with varying lignin content has been evaluated, by subjecting poplar, pine, and hay to solvent-based (water/ethanol/acetic acid) pretreatments and by preparing physical mixtures of pure microcrystalline cellulose with organosolv lignin. Lignin appeared to inhibit the activity of the hydrogenation catalyst, Raney-Ni, by hindering the formation of sugar alcohols in the presence as well as in the absence of the tungstate catalyst. However, lignin is not the root cause for the low EG yield typically obtained with untreated lignocellulose, as treated lignocellulose delivered high EG yields (30-35 wt %), irrespective of the lignin concentrations, which varied between 0 and 44 wt %, under identical demanding experimental conditions.