AbstractAutohydrolysis of Eucalyptus globulus was conducted at three different intensity levels typical for prehydrolysis kraft pulping as utilized for manufacturing dissolving pulp grades. The objective was to establish for the autohydrolysis process a detailed mass balance comprising the chemical composition of all three phases: the autohydrolysate, the released gas, and the solid residue. Carbohydrate determination involved both acid methanolysis combined with gas chromatography (GC) and sulfuric acid total hydrolysis with high performance anion exchange chromatography (HPAEC) coupled with pulsed amperometric detection (PAD); this allowed reliable quantification of neutral as well as acidic sugar units in cellulosic and non-cellulosic polysaccharides. Uronic acids present in the Eucalyptus globulus wood were progressively degraded through decarboxylation, leading to substantial carbon dioxide formation. The degree of acetylation of xylan remaining in the wood residue was clearly reduced, while the amount of bound acetyl groups in dissolved xylo-oligosaccharides (XOS) stayed relatively constant as a function of autohydrolysis intensity. The bulk of the lignin that was dissolved during autohydrolysis could be attributed to the acid-soluble lignin content of the wood. Only small amounts of Klason lignin were dissolved.