Hybrid poplar (Populus nigra x P. maximowiczii) chips were pretreated using an organosolv ethanol process. The effect of pretreatment conditions (temperature, time, catalyst, and ethanol concentration) on the substrate characteristics, including fiber size, crystallinity, and degree of polymerization of cellulose, was investigated using an experimental matrix designed with response surface methodology. The conditions ranged 155-205 degrees C, 26-94 min, 0.83-1.67% catalyst (H2SO4) on oven-dry wood chip (w/w), and 25-75% ethanol concentration (v/v). The results indicated that the substrate characteristics are controllable and predictable. Desirable substrates can be prepared by fine-tuning the processing parameters. The regression models developed, allowed the quantitative prediction of the substrate characteristics from the pretreatment conditions used.