Lignin is a complex phenylpropanoid polymer derived from enzyme mediated radical coupling of coniferyl, sinapyl and coumaryl alcohols with main functions to impart strength to plant cell wall, transport water and provide defense against pathogens. Physical, chemical and biological degradation of cellulose and hemicellulose is inversely proportional to the amount of lignification. Lignin therefore confers biomass recalcitrance and necessitates pretreatment steps, which are costly and must be improved if bioenergy from biomass is to be realized. New emerging pretreatment techniques employing ionic liquids have shown great promise and are very effective in breaking inter and intramolecular hydrogen bonding in cellulose microfibrils, thereby making cellulose amorphous and enhancing saccharification in both model cellulose and biomass. However, detailed understanding and insight into the extent of lignin depolymerization during pretreatment processes and the influence of degree of lignification, lignin chemical composition, and chemical association between lignin and hemicellulose are lacking. Towards the goal of gaining molecular level understanding of lignin signatures and their influence on depolymerization via pretreatment, we are utilizing light scattering, spectroscopy, chromatography and modified wet chemistry techniques. We aim to gain knowledge for attaining optimized pretreatment conditions, aiding efforts on altering chemistry in bioenergy crops, and perhaps selective depolymerization of celluloses or lignin. We will present results on our ongoing experiments on potential bioenergy crops with varied  dgree of lignification and lignin composition characterized before and after pretreatment. In addition, disruption of lignin-hemicellulose association during pretreatment will be discussed.