Hydrolysis is the process to break down the pretreated cellulosic molecules into cellobiose, which is then further converted to simple sugar, such as glucose molecules and short chains. Hydrolysis can be carried out biologically through enzymatic reaction or chemically using acid.

20.3.2.1 Saccharification by Enzymatic Processes

One of the basic methods of hydrolysis is enzymatic hydrolysis. Enzymatic hydro­lysis occurs when enzymes are exposed to the pretreated biomass to decompose the cellulosic materials into simple sugars. Cellulose in nature is mostly decomposed by cellulolytic fungi and bacteria enzymatically. Some of the microbes that can produce cellulose enzyme are Trichoderma reesei, Trichoderma viride, and Aspergillus niger.

Enzymatic degradation of cellulose to fermentable sugar is generally accom­plished by synergistic action of high specific cellulose. This group includes at least 15 protein families and some subfamilies (Rabinovich et al. 2002). Enzymatic deg­radation of cellulose to glucose is generally accomplished by synergistic action of at least three major classes of enzyme: endocellulase, exocellulase, and p-glucosidase. These enzymes are usually called together cellulose or cellulolytic enzymes (Wyman 1996). To work, enzymes must obtain access to the molecules to be hydro­lyzed. Therefore, pretreatment process is needed to break the crystalline structure of the lignocellulose and remove the lignin to expose the cellulose and hemicellulose molecules.

The enzymes digest the lignin surface yielding cellulose. The endocellulase and exocellulase digest the cellulose into polysaccharide molecules. The polysaccharide molecules are then digested by the p-glucosidase yielding the final glucose product (Klass 2008). The reaction occurs around 40-50 °C and at a pH of about 5. Below, the figure demonstrates how the reaction path occurs.

However, enzymatic hydrolysis can be problematic. The hydrolysis products (glucose and cellulose chains) inhibit the ability for enzymes to convert cellulose to glucose. As more products are formed, the enzymes become more inhibited by the excess glucose present. This ultimately slows down the hydrolysis process yielding low levels of usable hydrolysis product (D’amore 1991). Another disad­vantage of using enzyme is the high enzyme cost. The enzyme cost takes a part as much as 53-65 % of total chemical cost, while the chemical cost is around 30 % of the total cost.

Enzymatic hydrolysis of lignocellulose is facing a number of obstacles that diminish the enzyme performance. Recently, although enzyme price has decreased due to intensive research by giant enzyme producers, such as Novozymes and Genencor, minimization of enzyme loading is still needed in order to reduce the production cost. Thus affecting the time needed to complete the enzymatic hydrolysis process. Furthermore, high substrate concentration increases the problem of product inhibition which also results in lower performance of the enzymes. Then, lignin presence in the substrate, which shields the cellulose chains and absorbs the enzymes, is also a major obstacle for efficient hydrolysis. Moreover, the activity of enzymes could be lost due to denaturation or degradation.