**2.3 Biological pretreatment**

Biological pretreatment is based in the extracellular enzymes released by microorganisms in which enzymes degrade the noncellulosic components of the fiber surface. Biological pretreatment of fiber offers relevant advantages, such as low chemical and energy use that make it eco-friendly [25]. A great variety of microorganisms exists in nature, they are able to hydrolyze lignin, being the fungi the most studied [3]. Basidiomycetes white-rot fungi are responsible for lignin degradation in nature; they can break down not only lignin but also hemicellulose and cellulose. It has been reported that these microorganisms degrade lignin in a selective way that is able to offer potential biotechnological application [26]. However, recent studies have shown that many bacteria are able to break down lignin [27]. Likewise, enzymes have an enormous potential to be used for lignin valorization.

#### *2.3.1 Fungal lignin degradation*

The breaking down of lignin by fungi has been reported mainly for white-rot fungi due to their highly efficient enzymatic system. White-rot fungi are able to degrade lignin in such an efficiently and selectively way that gives them utility in the industry. These fungi have been applied by different industries such as paper, biofuels, and biorefinery for delignifying biomass [28]. According to the selected strain, it is possible to obtain 20–100% for lignin removal. Black liquor from a pulp and paper mill, treated with the fungi *Pleurotus ostreatus,* reduced 70% its lignin content [29]. Sugarcane bagasse treated with *Lentinula edodes* and *P. ostreatus* presented, after the treatment, 87 and 85% of lignin, respectively [30]. Biological pretreatment of bamboo culms with *Punctularia* sp. Strain TUFC20056 showed more than 50% on lignin degradation [31]. High ligninolytic capabilities have

**41**

*Getting Environmentally Friendly and High Added-Value Products from Lignocellulosic Waste*

been found in the fungi *Polyporus brumalis* using wheat straw as substrate [32]. The fungal lignin degradation is based in an oxidative system. The oxidative and ligninolytic system is based in extracellular enzymes, which break down lignins and open phenyl rings; these enzymes are divided into two families: polyphenol oxidases (laccases) and lignin-modifying heme-containing peroxidases (LMPs); this second family comprises: lignin peroxidases (LiP), manganese peroxidases (MnP),

Laccases use molecular oxygen to oxidize aromatic and nonaromatic compounds, such as phenols, arylamines, anilines, thiols, and lignins [34]. The oxidation leads to the constitution of free radicals that act as intermediate for the enzymatic reactions. Likewise, these mediators can react with others high redox potential compounds and mediate nonenzymatic reactions [26]. White-Rot fungi are mainly reported to produce laccases such as, *Phlebia radiata*, *P. ostreatus,* and *Trametes versicolor* [35]. Although this enzyme is generally found in fungi, it has been found in bacteria as well, such as *Streptomyces lavendulae*, *S. cyaneus*, and *Marinomonas mediterranea* [36]. Laccases present an enormous potential because they work efficiently on a broad range of substrates with applications on paper industries, biosensors (identifying morphine or codeine), food industries, textile

industries, soil bioremediation, and in the production of polymers [37].

LMPs belong to class II peroxidases, named plant, and fungal peroxidases, which contain protoporphyrin IX as a prosthetic group [38]. LiP enzymes oxidize different phenolic aromatic compounds and nonphenolic lignin compounds due to the fact that they are not very specific to their substrates [39]. LiP enzymes have been found only in a few white-rot fungi such as the genera: *Bjerkandera*, *Phanerochaete*, *Phlebia*, and *Trametes* [40–42]. The most common peroxidases found in white-rot fungi and other litter-decomposing fungi are the glycoproteins MnP [43]. The MnP glycoproteins catalyze the oxidation of Mn (II) to Mn (III), which is released in complex with oxalate or others chelators [44]. MnP enzymes are found in white-rot wood and litter-decomposing fungi such as *Dichomitus squalens*, *Agaricus bisporus*, and *Agrocybe praecox* [45]. VP enzymes present molecular similarities to LiP and MnP, oxidizing substrates as LiP and Mn2+ with a similar catalytic site to MnP [38]. VP enzyme has been found in white-rot fungal in the genera *Pleurotus* and *Bjerkandera* [46]. A variety of low molecular weight aromatic compounds are obtained from fungal lignin degradation, such as, guaiacol, coniferyl alcohol, p-coumarate, ferulate, protocatechuate, p-hydroxybenzoate, and vanillate [47]. The resulting liquor can be used by bacteria that can metabolize lignin-derived aromatics compounds [48].

It has been reported that bacteria are able to degrade lignin through a complex

of enzymes, such as extracellular peroxidases, Dye-decolorizing peroxidases (DyPs), and laccases. Among the reported bacterial genus, we found *Rhodococcus*, *Pseudomonas*, *Streptomyces, Novosphingobium*, and *Bacillus* [49]. The bacteria *S. viridosporus* and *N. autotrophica* were able to degrade lignin through extracellular peroxidases, whereas *P. putida*, *Rhodococcus RHA1*, and *Rhodococcus* sp. were active in hydrogen peroxide absence suggesting the presence of extracellular laccases [50]. DyP peroxidases are able to oxidize lignin, aromatic dye, and other phenolic compounds [51]. In spite of finding at first the DyP peroxidases in fungi, recent studies have shown that these enzymes are prominent in bacteria [52]. Bioinformatic analysis showed that *R. jostii* sp. presents two peroxidases members of the DyP peroxidase family, and the deletion mutant gene assay in these genes showed reduced lignin degradation [53]. Bacterial laccases have showed high tolerance to temperature, salt, and acid/alkaline conditions, which make them valuable in the industry,

*DOI: http://dx.doi.org/10.5772/intechopen.93645*

and versatile peroxidases (VP) [33].

*2.3.2 Bacterial lignin degradation*

#### *Getting Environmentally Friendly and High Added-Value Products from Lignocellulosic Waste DOI: http://dx.doi.org/10.5772/intechopen.93645*

been found in the fungi *Polyporus brumalis* using wheat straw as substrate [32]. The fungal lignin degradation is based in an oxidative system. The oxidative and ligninolytic system is based in extracellular enzymes, which break down lignins and open phenyl rings; these enzymes are divided into two families: polyphenol oxidases (laccases) and lignin-modifying heme-containing peroxidases (LMPs); this second family comprises: lignin peroxidases (LiP), manganese peroxidases (MnP), and versatile peroxidases (VP) [33].

Laccases use molecular oxygen to oxidize aromatic and nonaromatic compounds, such as phenols, arylamines, anilines, thiols, and lignins [34]. The oxidation leads to the constitution of free radicals that act as intermediate for the enzymatic reactions. Likewise, these mediators can react with others high redox potential compounds and mediate nonenzymatic reactions [26]. White-Rot fungi are mainly reported to produce laccases such as, *Phlebia radiata*, *P. ostreatus,* and *Trametes versicolor* [35]. Although this enzyme is generally found in fungi, it has been found in bacteria as well, such as *Streptomyces lavendulae*, *S. cyaneus*, and *Marinomonas mediterranea* [36]. Laccases present an enormous potential because they work efficiently on a broad range of substrates with applications on paper industries, biosensors (identifying morphine or codeine), food industries, textile industries, soil bioremediation, and in the production of polymers [37].

LMPs belong to class II peroxidases, named plant, and fungal peroxidases, which contain protoporphyrin IX as a prosthetic group [38]. LiP enzymes oxidize different phenolic aromatic compounds and nonphenolic lignin compounds due to the fact that they are not very specific to their substrates [39]. LiP enzymes have been found only in a few white-rot fungi such as the genera: *Bjerkandera*, *Phanerochaete*, *Phlebia*, and *Trametes* [40–42]. The most common peroxidases found in white-rot fungi and other litter-decomposing fungi are the glycoproteins MnP [43]. The MnP glycoproteins catalyze the oxidation of Mn (II) to Mn (III), which is released in complex with oxalate or others chelators [44]. MnP enzymes are found in white-rot wood and litter-decomposing fungi such as *Dichomitus squalens*, *Agaricus bisporus*, and *Agrocybe praecox* [45]. VP enzymes present molecular similarities to LiP and MnP, oxidizing substrates as LiP and Mn2+ with a similar catalytic site to MnP [38]. VP enzyme has been found in white-rot fungal in the genera *Pleurotus* and *Bjerkandera* [46]. A variety of low molecular weight aromatic compounds are obtained from fungal lignin degradation, such as, guaiacol, coniferyl alcohol, p-coumarate, ferulate, protocatechuate, p-hydroxybenzoate, and vanillate [47]. The resulting liquor can be used by bacteria that can metabolize lignin-derived aromatics compounds [48].
