**2. Physical methods**

Pore size and surface area of lignocellulosic biomass can be increased, whereas crystallinity and degree of polymerization of cellulose can be decreased with the

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0.5 mm particle size [25] (**Figures 1** and **2**).

*Different Pretreatment Methods of Lignocellulosic Biomass for Use in Biofuel Production*

application of physical methods. Physical pretreatments include milling, sonication,

On the inherent ultrastructure of cellulose and degree of crystallinity, milling can be performed to render lignocelluloses more amenable to cellulases. Cellulases are enzyme that catalyze cellulose, but for the catalysis and best results, the substrate availability needs to be enhanced for optimized functioning of the enzymes. Before the subjection of the LCW to enzymatic hydrolysis, milling and size reduction of the lignocellulosic matter should be performed. Milling process has several types like ball milling, colloid milling, vibro-energy milling, hammer milling, and two-roll milling. For wet material, colloid mill, dissolver, and fibrillator are suitable, whereas for dry materials hammer mill, extruder, cryogenic mill, and roller mill are used. For both wet and dry material, ball milling can be used. For waster paper, hammer milling is the most suitable pretreatment option. Enzymatic degradation can be improved by milling as it reduces the degree of crystallinity and material size. Up to 0.2 mm reduction in particle size can be seen by milling and grinding. Reduction in particle size of biomass can be achieved up to a certain limit; beyond that limit reduction in particle size does not effect in the pretreatment procedure. Corn stover with small particle size, i.e., from 53 to 75 μm, is more productive as compared to large particle size corn stover ranging from 475 to 710 μm. The difference in particle size shows that productivity can significantly affect the pretreatment process. Ball milling causes a massive drop in crystallinity index from 4.9 to 74.2% which makes this process more suitable for saccharification of straw at mild hydrolytic conditions with more production of fermentable sugars [12, 16–18]. For better results of hydrolysis, milling can be used in combination with enzymatic hydrolysis. Mechanical action, mass transfer, and enzymatic hydrolysis can be achieved at the same time when two methods are combined. A number of ball beads in bill mill reactor play a crucial role in the α-cellulose hydrolysis, as less enzyme loading is required, and 100% rate of hydrolysis can be achieved in comparison to pretreatment of biomass that is carried without the use of milling procedure. Highest hydrolysis rate with high yield of reducing sugar was obtained when rice straw was put into fluidized bed opposed jet mill for fine grinding after cutting, steam explosion, and pulverization. For pretreatment of biomass, ball milling is an expensive option in terms of energy consumption, which is a huge disadvantage at industrial scale. Also, incapability of milling for removing lignin makes it a less suitable option as enzyme accessibility to the substrate is reduced in the presence of lignin. Reduction in crystallinity, degree of polymerization, and increase in surface area can be effected by the type of biomass, type of milling used for pretreatment, and duration of the milling process [19–21]. For improving digestibility and reducing crystallinity, vibratory ball milling is very effective. Low energy consumption has an important advantage of using wet disk milling which produces fibers that improve hydrolysis of cellulose, whereas hammer milling produces finer bundles. Due to this reason milling is not preferred when wet disk milling is available [22, 23]. Other study results of conventional ball and disk milling are compared. With the use of conventional ball milling, maximum yields of xylose and glucose were obtained, i.e., 54.3 and 89.4%, respectively [24]. Wet milling produces less yield, but it has the advantage of not producing inhibitors and very low energy consuming capability. An increase of 110% in enzymatic hydrolysis was achieved when wet milling was combined with alkaline pretreatment. Optimum parameters for wet milling pretreatment of corn stover were 10 mm diameter 20 steel balls, 1:10 solid-to-liquid ratio, 350 rpm/min speed, and

*DOI: http://dx.doi.org/10.5772/intechopen.84995*

**2.1 Milling**

mechanical extrusion, ozonolysis, and pyrolysis.

application of physical methods. Physical pretreatments include milling, sonication, mechanical extrusion, ozonolysis, and pyrolysis.
