In lignocellulosic biorefinery processes involving enzyme catalysed reactions it is a challenge that enzyme inhibiting compounds are generated and liberated during pretreatment of the biomass. In this study the contribution to cellulase inhibition from xylooligosaccharides and newly discovered oligophenolic compounds from pilot scale pretreated wheat straw was assessed at two different pretreatment severities. An increase in severity of the pretreatment led to more oligophenol compounds and in turn the total overall cellulase inhibition increased. When the xylooligosaccharides were enzymatically degraded prior to cellulose hydrolysis, a relief in cellulase inhibition was observed, but some inhibition remained, suggesting that other components also played a role in inhibition. We propose that these components include dipentoses with bicyclic moieties and feruloylated tripentoses, because LC-MS/MS analysis revealed the presence of these components in the liquid from hydrothermal pretreated wheat straw after enzymatic treatment. The reaction mechanisms for synthesis of the new dipentoses having hydroxylated oxane bicyclic residues are considered and they are proposed to be formed as reaction products from either xylose or glucose reacting with glyceraldehyde during pretreatment. The data show that the main cellulase inhibition from hydrothermally pretreated wheat straw liquors is due to xylooligosaccharides followed by oligophenolic compounds and the newly discovered dipentose with bicyclic moieties and feruloylated tripentoses. The relative amounts and hence contribution to inhibition from each class of compounds changes with severity of the pretreatment.