The use of enzymes for the pretreatment of biomass feedstocks can significantly reduce the capital cost, under the condition that enzymes are produced by microorgan­isms during the process of fermentation, also known as simultaneous saccharification

and fermentation (SSF) (Olofsson et al. 2008). However, the major drawbacks of SSF are the need to find optimal conditions of temperature and pH for both the enzymatic hydrolysis and the fermentation, and the difficulty to recycle the fermenting organism and the enzymes (Olofsson et al. 2008). Nevertheless, the application of enzymes can facilitate the fast, efficient, and cheap conversion of cellulose to glucose. Enzymatic hydrolysis can give higher yields of sugars in contrary to acid hydrolysis. However, the enzyme system for hydrolyzing lignocellulose is quite complex and involves the cellulase system as shown in Table 4. The efficacy of the enzyme systems depends on various factors that should be overcome to achieve the maximum yields of glucose. The key barriers that impede the action of enzyme system are as follows: (1) unreac­tive nature of crystalline cellulose; (2) the presence of lignin-blocking reactive sites; (3) low substrate surface area; (4) low rates of hydrolysis; (5) substrate and product inhibition; and (6) enzyme denaturation.

In order to develop an effective enzymatic hydrolysis process, it is important that inhibitors that impact the enzyme activity are removed (Taherzadeh and Karimi 2007). Another issue that requires attention is the cost reduction of the enzymes, which can be achieved probably by recycling the enzyme or by producing micro­bial enzymes during the SSF process. Recycling of enzymes can be achieved by repeated batch hydrolysis of feedstocks and immobilization of enzymes on an inert material (Das et al. 2011). The application of immobilized enzyme enables easy post­hydrolysis separation of the enzyme from the reaction mixture (Das et al. 2011). The advantage of enzyme immobilization is that it ensures the enzyme structure and conformation is preserved in addition to imparting improved thermostability. Enzyme modifications and active site mutations could possibly provide much effec­tive enzymes with high rates of hydrolysis, reusability, and resistance to denaturation. Modified/novel enzymes have the potential to reduce the cost of enzymatic hydrolysis.