The fuel properties of fast pyrolysis bio-oils differ significantly from those of fossil fuels. As transportation fuel, bio-oil is not suitable without upgrading because of its relatively low energy content, high water content, acidity, and poor storage stability. Upgrading of bio-oil has usually been done by treating the whole oil in a reactor. The problem with this treatment is that pyrolysis oil is a mixture of different compound groups, which all need different conditions and catalysts to react in a desirable way. Therefore, an efficient fractionation of bio-oil before upgrading may be a more efficient way of producing liquid fuels than treating the whole oil. In this work, the target was to compare two industrially relevant fractionation concepts. In the first concept, most of the water was removed during liquid recovery by adjusting the scrubber temperature. When the scrubber temperature was increased from 36 to 66 °C, the water content in the bio-oil decreased from 24 to 7 wt %. In the second concept, fast pyrolysis was carried out with wet feedstock. This would reduce the drying cost in the plant. By this means, a spontaneous phase separation was generated after liquid condensation. In the experiments, the moisture content of the raw material was increased up to 25 wt %, but even with this moisture content, the oily bottom phase still contained 22 wt % watersoluble compounds. However, if the target is to produce transportation fuels from bio-oil, fractionation by phase separation is a better concept for dividing the bio-oil into different compound groups.