Attempts to correlate the physical and chemical properties of biomass to its susceptibility to enzyme digestion are often inconclusive or contradictory depending on variables such as the type of substrate, the pretreatment conditions and measurement techniques. In this study, we present a direct method for measuring the key factors governing cellulose digestibility in a biomass sample by directly probing cellulase binding and activity using a purified cellobiohydrolase (Cel7A) from Trichoderma reesei. Fluorescence-labeled T. reesei Cel7A was used to assay pretreated corn stover samples and pure cellulosic substrates to identify barriers to accessibility by this important component of cellulase preparations. The results showed cellulose conversion improved when T. reesei Cel7A bound in higher concentrations, indicating that the enzyme had greater access to the substrate. Factors such as the pretreatment severity, drying after pretreatment, and cellulose crystallinity were found to directly impact enzyme accessibility. This study provides direct evidence to support the notion that the best pretreatment schemes for rendering biomass more digestible to cellobiohydrolase enzymes are those that improve access to the cellulose in biomass cell walls, as well as those able to reduce the crystallinity of cell wall cellulose.

The crystallinity index of cellulose is an important parameter to establish because of the effect this property has on the utilization of cellulose as a material and as a feedstock for biofuels production. However, it has been found that the crystallinity index varies significantly depending on the choice of instrument and data analysis technique applied to the measurement. We introduce in this study a simple and straightforward method to evaluate the crystallinity index of cellulose. This novel method was developed using solid state 13 C NMR and subtraction of the spectrum of a standard amorphous cellulose. The crystallinity indexes of twelve different celluloses were measured and the values from this method were compared with the values obtained by other existing methods, including methods based on X-ray diffraction. An interesting observation was that the hydration of the celluloses increased their crystallinity indexes by about 5%, suggesting that addition of water increased cellulose order for all the cellulose samples studied.

Enzyme accessibility has been proposed as a limiting factor in the enzymatic conversion of the cellulose in biomass to glucose. Prior work has shown a strong correlation between porosity, measured as the change in the volume of pores accessible to a cellulase-sized molecule, and the initial digestibility of biomass pretreated by various methods. The goal of this work was to determine if porosity was one of the factors governing the overall enzymatic digestibility of the cellulose in dilute acid pretreated biomass. The porosity of wet pretreated corn stover was determined using the methods of solute exclusion and 1H nuclear magnetic resonance (NMR) thermoporometry. The solute exclusion method identified differences in the accessible pore volume of the pretreated samples compared to untreated corn stover; however, only very small differences in porosity were observed among samples pretreated with a range of severities, giving ethanol yields from 70 to 96%. No correlation was found between the volume accessible to an enzyme-sized molecule (diameter estimated to be 51 A) and the digestibility of the cellulose in dilute acid pretreated corn stover. 1H NMR thermoporometry was used to measure the amount of water in pores ranging from 20 to 200 A. As was the case for the solute exclusion method, a difference was observed in the pore volume of untreated and acid pretreated corn stover, but no significant differences in pore volume were measured for the different pretreated samples.

It has previously been shown that the improved digestibility of dilute acid pretreated corn stover is at least partially due to the removal of xylan and the consequent increase in accessibility of the cellulose to cellobiohydrolase enzymes. We now report on the impact that lignin removal has on the accessibility and digestibility of dilute acid pretreated corn stover. Samples of corn stover were subjected to dilute sulfuric acid pretreatment with and without simultaneous (partial) lignin removal. In addition, some samples were completely delignified after the pretreatment step using acidified sodium chlorite. The accessibility and digestibility of the samples were tested using a fluorescence-labeled cellobiohydrolase (Trichoderma reesei Cel7A) purified from a commercial cellulase preparation. Partial delignification of corn stover during dilute acid pretreatment was shown to improve cellulose digestibility by T. reesei Cel7A; however, decreasing the lignin content below 5% (g g -1 ) by treatment with acidified sodium chlorite resulted in a dramatic reduction in cellulose digestibility. Importantly, this effect was found to be enhanced in samples with lower xylan contents suggesting that the near complete removal of xylan and lignin may cause aggregation of the cellulose microfibrils resulting in decreased cellulase accessibility.

It has been proven that it is possible to improve the performance of water flooding (WF hereafter) by chemically altering the brine/water composition. There are a number of compounds that can be added for this purpose and CO2 is one of them. Carbonated water flooding (CWF hereafter) is to inject CO2 saturated (or nearly saturated) water into reservoirs as the displacing fluid. CO2 stays in the water phase first and migrates into the oil phase afterwards, without forming an individual CO2 rich phase. Such mass transfer of CO2 from water into oil is substantial due to the fact that under the same pressure and temperature conditions, CO2 is more soluble in oil than in water. With the dissolution of CO2, oil viscosity is reduced, which makes mobility ratio between water and oil more favorable in the contacted zone, and oil volume expands (swelling effects), which increases the relative permeability of oil. Both effects result in improved ultimate recovery over conventional WF. Since miscibility is not sought during the CWF process, it has less restrictive requirements of reservoir conditions and oil types. In addition, ease of separation of CO2 water mixture at the production wells and less gas handling make CWF relatively easier to implement in fields with ongoing WF and/or in future candidates. Moreover, at the end of CWF cycle, it is still possible to implement other tertiary flooding techniques as in the case of conventional WF. CWF is an improved oil recovery (IOR hereafter) method that has been tested in the field [6][15][16][26][35]. Pilot programs and field trials were conducted in 1950's. Lab scale experiments were also active between 60's and 80's. During the past a few decades, however, little work was done on the topic. Literature data in this area are inconclusive in several aspects, for example the large variations of the incremental recovery and experimental conditions that can be represented in terms of dimensionless scaling factors (that control the displacement performance). This paper summarizes the results of a series of CWF experiments conducted using a specifically designed core flooding apparatus at our labs. These experiments consist of the Phase I study of a project designed to obtain a comprehensive understanding of CWF's displacement mechanism as well as the impact of several pre-identified sensitivity parameters, such as injection rate, salinity, etc. Eight sand pack experiments (divided into four sub groups) were completed under the same pressure, temperature, and CO2 saturation level, yet with different injection rates. The main contributions of this study are: Overall, CWF results in better oil recovery than its WF counterpart with improvement ranges from 35% (high speed, 15 PV/day) to 3% (low speed, 1 PV/day) of STOIIP, using a medium viscosity crude oil. Cross-comparison and interpretation of current and historical experimental results in terms of identification of flow regimes utilizing dimensionless numbers, such as gravity number, capillary number, etc. Identification of the optimal flow rate and the range of incremental recovery over WF. Providing new data for the literature.