**5.2 Enzymatic saccharification**

The pretreatment method aims at facilitating maximum saccharification of cellulose and hemicellulose by enzymatic hydrolysis. In this process, the cellulose and hemicellulose present in the biomass will be hydrolysed by cellulase and hemicellulase enzyme produced from fungi into simple monomer sugar such as glucose and xylose. This monomer sugar is the main chemical platform for biofuel and other chemicals (**Figure 8**). Previous research obtained the cellulose from untreated biomass and biofiber upon enzymatic hydrolysis can yield not more than 15–25% glucose due to the recalcitrance [76]. Most pretreatment methods have some disadvantages in terms of cost, recovery, secondary pollution, and formation of intermediate compounds that will inhibit enzymatic hydrolysis, but implementation of laser, microwave, and electron beam irradiation have become more attractive because of its fast and effective result during experimentation [77].

Biomass pretreated with electron beam irradiation (EBI) enhance enzymatic saccharification by decreasing the crystallinity and molecular weight and simultaneously increase the surface area [36, 68]. Irradiation induces a chain-cleavage mechanism by depolymerizing the poly‐ meric material [78]. Higher cellulose content was found in chemical-irradiated pre-treated oil palm empty fruit bunch (OPEFB) than untreated OPEFB. Higher cellulose content can produce higher glucose, hemicelluloses content can be converted to xylosa, while lignin can produce derivatives compound of phenol [36]. The effectiveness of EBI treatment also depends on the nature of biomass with respect to energy delivered, and sources and concentration of enzymes used [77]. The earliest study by Kumakura and Kaetsu [79] found the pre-irradiation (dosage 107 rad) with presence of chlorine yields six times higher reducing sugar than its absence with subsequent enzymatic hydrolysis on rice straw. Then Ardica et al. [80] used gamma-ray irradiation (doses range from 1 kGy to 1000 kGy) on wood chips, kapok, papers, hays, and grain straw to enhance the enzymatic hydrolysis. Combined pretreatment of gamma-ray and

Normally, the structure of lignin consists of guaiacyl propane units (G) and syringyl propane units (S) containing one and two methoxy groups. It is known that the presence of guaiacyl propane could restrict the swelling of biomass [73, 74]. These chemical structures can be identified by FT-IR spectra with frequencies in the region of 1509, 1464, and 1422 cm−1. Reduction of spectra in this region indicated that most of the lignin in OPT and OPF have been removed from the biomass during the pretreatment process. Removal of lignin in the biomass after irradiation gives a better access for enzyme to attack cellulose and hemicellulose.

The FT-IR analysis of the irradiation pretreatment on biomass also indicated that significant changes on absorbance was observed at 1732 cm−1 and 3300 cm−1, attributed to the vibration of hydrogen bonded OH-group. Liu et al. [75] reported a shifting and reduction of band 2899 cm −1, indicating to the disruption of biomass resulting from the C-H shifting vibration. The study also found that high-energy irradiation pretreatment could interrupt and destroy the intramolecular and inter molecular hydrogen bond in the cellulose. The degradation of cellulose generated carbonyl group could be determined at band at 1603 cm−1. Apart of this region, the shifting of band region between 1164 cm−1, 1112 cm−1, and 1058 cm−1 attributed to the vibration

The pretreatment method aims at facilitating maximum saccharification of cellulose and hemicellulose by enzymatic hydrolysis. In this process, the cellulose and hemicellulose present in the biomass will be hydrolysed by cellulase and hemicellulase enzyme produced from fungi into simple monomer sugar such as glucose and xylose. This monomer sugar is the main chemical platform for biofuel and other chemicals (**Figure 8**). Previous research obtained the cellulose from untreated biomass and biofiber upon enzymatic hydrolysis can yield not more than 15–25% glucose due to the recalcitrance [76]. Most pretreatment methods have some disadvantages in terms of cost, recovery, secondary pollution, and formation of intermediate compounds that will inhibit enzymatic hydrolysis, but implementation of laser, microwave, and electron beam irradiation have become more attractive because of its fast and effective

Biomass pretreated with electron beam irradiation (EBI) enhance enzymatic saccharification by decreasing the crystallinity and molecular weight and simultaneously increase the surface area [36, 68]. Irradiation induces a chain-cleavage mechanism by depolymerizing the poly‐ meric material [78]. Higher cellulose content was found in chemical-irradiated pre-treated oil palm empty fruit bunch (OPEFB) than untreated OPEFB. Higher cellulose content can produce higher glucose, hemicelluloses content can be converted to xylosa, while lignin can produce derivatives compound of phenol [36]. The effectiveness of EBI treatment also depends on the nature of biomass with respect to energy delivered, and sources and concentration of enzymes used [77]. The earliest study by Kumakura and Kaetsu [79] found the pre-irradiation (dosage 107 rad) with presence of chlorine yields six times higher reducing sugar than its absence with subsequent enzymatic hydrolysis on rice straw. Then Ardica et al. [80] used gamma-ray irradiation (doses range from 1 kGy to 1000 kGy) on wood chips, kapok, papers, hays, and grain straw to enhance the enzymatic hydrolysis. Combined pretreatment of gamma-ray and

of C-O-C of cellulose.

344 Radiation Effects in Materials

**5.2 Enzymatic saccharification**

result during experimentation [77].

**Figure 8.** Enzymatic saccharification of cellulose and hemicellulose to monomer sugars by cellulase and hemicellulose enzyme.

diluted acid on poplar bark biomass observed a drastic increased in reducing sugar yield from 56.1 to 83.1% compared to gamma-ray pretreatment alone [81]. **Table 4** shows previous study using electron beam irradiation with various types of biomass.

Zhu et al. [32] reported the rice straw pretreated with microwave-alkali method obtained higher hydrolysis rate than alkaline pretreated alone. The amount of glucose obtained from the enzymatic hydrolysis using *Trichoderma reesei* cellulase was higher (24.8 gl−1), and lower for xylose (2.6 gl−1) concentration for microwave/alkali pretreated rice straw which is more suitable for subsequent fermentation process to produce biofuel. The microwave-alkali assist irradiation has been proven to remove more hemicellulose and lignin, and simultaneously increasing enzyme accessibility [32]. In 2006, they have presented comparison between three techniques for the enzymatic hydrolysis of rice straw by pre-treating them with microwave/ alkali, microwave/acid/alkali, and microwave/acid/alkali/H2O2 treatment. The result shows that the rice straw pretreated with microwave/acid/alkali/H2O2 treatment had the highest hydrolysis rate and glucose content in the hydrolysate. Furthermore, recovery of xylose content could be recovered compared to microwave/alkali treatment [83]. [84] using micro‐ wave-assisted alkali treatment. They have presented at optimal condition of 190°C, 50g/l solid content, and 30 min treatment time. The sugar yield from the combined treatment and hydrolysis was 58.7g/100g biomass which is equivalent to 99% of potential maximum sugars. This study was further investigated by using scanning electron microscope, and showed the advantage of microwave over conventional heating was due to the disruption of recalcitrant structures of lignocellulose. In conclusion, microwave/chemical pretreatment is more effective than conventional heating.


a, pretreatment added with diluted sulfuric acid.

b, pretreatment added with water soaking-based EBI.

na, not available.

**Table 4.** Summarization of previous research using EBI pretreatment on various types of biomass and biofiber.
