Application of advanced integrated imaging methods in the field of biomass degradation for biofuel production

In order to better understand and focus on the biological systems related to biofuel production, especially during the process of biomass pretreatment and enzyme hydrolysis, novel and integrated advanced imaging techniques are being developed at Joint Bio-Energy Institute (JBEI).  Recently, certain ionic liquids (ILs) have been used during pretreatment to mitigate biomass recalcitrance to enzymatic hydrolysis and/or fermentation during the process of biofuel production. Although there are several reports about the reduction of the surface crystallinity after ILs pretreatment, the overall mechanism of ILs pretreatment is still not fully understood. Meanwhile the enzymatic hydrolysis by thermostable and/or IL-tolerant enzymes is still not clear for their work mechanistically on IL pretreated biomass. In order to better understand the saccharification of cellulosic biomass to enhance the efficiency of fermentable sugar production, it is critical to gain better understanding of the saccharification and IL pretreatment process with high resolution imaging. To meet this need, we have combined optical-AFM (atomic force microscope) technique for high resolution imaging in liquid with force spectroscopy to gain mechanistic insight of pretreatment process and follow the enzymatic hydrolysis of multiple biomass feed-stocks in real-time. The in-solution AFM is used to obtain high-resolution and real-time images during the ILs pretreatment and the enzymatic hydrolysis. Combining these results together with other advanced optical systems such as the confocal fluorescent microscope, ramam spectroscopy and XRD, we have realized a multi-scale imaging analytical capability that provides robust chemical and crystal structure information in the area of ionic liquid pretreatment and enzymatic hydrolysis.