Three hydrotreated bio-oils with different oxygen contents (8.2, 4.9, and 0.4 w/w) were distilled to produce light, naphtha, jet, diesel, and gas oil boiling range fractions that were characterized for oxygen-containing species by a variety of analytical methods. The bio-oils were originally generated from lignocellulosic biomass in an entrained-flow fast pyrolysis reactor. Analyses included elemental composition, carbon type distribution by C-13 nuclear magnetic resonance, acid number, gas chromatography/mass spectroscopy, volatile organic acids by liquid chromatography, and carbonyl compounds by 2,4-dinitrophenylhydrazine derivatization and liquid chromatography. Acid number titrations employed an improved titrant electrode combination with faster response that allowed the detection of multiple end points in many samples and allowed for acid values attributable to carboxylic acids and to phenols to be distinguished. The results of these analyses showed that the highest oxygen content bio-oil fractions contained oxygen as carboxylic acids, carbonyls, aryl ethers, phenols, and alcohols. Carboxylic acids and carbonyl compounds detected in this sample were concentrated in the light, naphtha, and jet fractions (<260 degrees C boiling point). The carboxylic acid content of all of the high oxygen content fractions was likely too high for these materials to be considered as fuel blendstocks, although the potential for blending with crude oil or refinery intermediate streams may exist for the diesel and gas oil fractions. The 4.9% oxygen sample contained, almost exclusively, phenolic compounds found to be present throughout the boiling range fractions, which imparted measurable acidity primarily in the light, naphtha, and jet fractions. Additional study is required to understand what levels of the weakly acidic phenols could be tolerated in a refinery feedstock. The diesel and gas oil fractions from this upgraded oil had low acidity but still contained 3-4 wt % oxygen present as phenols that could not be specifically identified. These materials appear to have excellent potential as refinery feedstocks and some potential for blending into finished fuels. Fractions from the lowest oxygen-content oil exhibited some phenolic acidity but generally contained very low levels of oxygen functional groups. These materials would likely be suitable as refinery feedstocks and potentially as fuel blend components. Paraffins, isoparaffins, olefins, naphthenes, and aromatics (PIONA) analysis of the light and naphtha fractions showed benzene contents of 0.5 and 0.4 vol % and predicted (research octane number (RON) + motor octane number (MON))/2 of 63 and 70, respectively.