3.3 Moisture content

Moisture content of the solid state fermentation is a critical aspect for fungal growth and activities. Lignin degradation is significantly influenced by this factor as it affects the growth and activities of the fungal. Increasing the moisture content enhances the nutrient transfer but reduces the porosity of the substrate and limits oxygen transfer [28]. However, insufficient water content in the substrates may cause deactivation of the fungi. Optimum moisture content depends upon the organism and the substrate used for SSF [30]. The range of moisture content of substrate for SSF using white-rot fungi is usually between 45 and 85% [29]. A study on the effect of moisture content for delignification of cotton stalks by Daedalea flavida MTCC 145 (DF-2) in SSF found that the highest ligninolytic enzyme

Substrate

48

Oil palm empty

P. ostreatus

P.

chrysosporium

> Oil palm empty

P. floridanus N.S

 59.40%

 31

 N.S 28

 +

• Lignin and cellulose were reduced by 0.03 and 5.0%, while

hemicellulose

•

Digestibility

 was improved by 4.5 times

 was increased by 4.4%

fruit bunches

Poplar wood

N.S, not specified.

Table 3.

Summary of recent publications

 on fungal

pretreatment.

T. velutina

100 v/w% N.S

28

 N.S 56

 N.S •

Delignification

cellulose were increased by 1.0 and 6.4%

 by 7.2%, whereas both

hemicellulose

 and

[27]

N.S

 67%

30

 N.S 21

 N.S •

Lignin, and 7.6%

> •

Reduction of lignin,

and 28.2%

hemicellulose

 and cellulose were 42.1, 27.7

[26]

hemicellulose

 and cellulose degradation

 at 51.9, 13.8

Biomass for Bioenergy - Recent Trends and Future Challenges

[25]

fruit bunches

Fungi

Inoculum

Moisture

T (°C) pH Time

species

conc.

content (%)

(days)

Nutrient

 Effect

References


fungi has various optimal temperature [33, 50]. White-rot basidiomycetes grow optimally at temperature between 25 and 30°C while most of white-rot ascomycetes fungi grow optimally at 39°C [5]. The metabolism of these fungi produces heat and develops temperature gradients in SSF media. The accumulated heat can lead to adverse effect on the fungal growth and their metabolic activity which leading to the denaturation of the key enzymes. From the studies on pretreatment of rice straw, ligninolytic activities by S. commune was found to be peaked at 30 and 35°C [40, 41]. Meanwhile the highest ligninases production by T. versicolor was reported at 40°C [51]. Adekunle et al. [33] reported in their study on the SSF of steamexploded cornstalk with T. versicolor that there was a direct correlation between the

Fungal Pretreatment of Lignocellulosic Materials DOI: http://dx.doi.org/10.5772/intechopen.84239

temperature and laccase production, with the highest laccase activity of

mum production (19,944 Ug<sup>1</sup> dry substrate), was found at 30°C [42].

3.5 pH

activity [5].

3.6 Aeration

3.7 Supplements

51

2677.16 U g<sup>1</sup> was produced at 28°C. The maximum production of laccase by T. giganteum AGHP (1.53 <sup>10</sup><sup>5</sup> Ug<sup>1</sup> of dry substrate) was obtained at 30°C [39]. This study also showed that lower temperature of 10 and 20°C are not suitable for the growth of fungi due to lower enzyme production. Similar result was obtained from the study on laccase production by Pseudolagarobasidium acaciicola LA 1, the opti-

pH is one of the prominent parameters in the cultivation of fungi and it is very problematic to control in SSF [52]. The initial pH of the medium influences the microbial growth and the production of ligninolytic enzyme. White-rot fungi grow well at pH 4–5, while substrate acidity decreases their growth. A study conducted on the isolation of laccase from a novel white-rot fungus Pseudolagarobasidium acaciicola LA 1 through SSF of Parthenium biomass showed that the isolated laccase was found to perform optimally at pH 4.5 and highly stable within the range of pH 4–7 for 24 h [42]. The effect of pH is important in the case of laccase production, and a small change in intracellular pH will result in a decrease of macromolecules synthesis. Patel and Gupte [39] reported that the maximal laccase production (1.27 <sup>10</sup><sup>5</sup> Ug<sup>1</sup> of dry substrate) by Tricholoma giganteum AGHP was achieved at pH 5.0. No increment in the production of enzyme was found at higher pH. This may be attributed to the poor mycelial growth at an elevated pH which may restrict the laccase production. It was reported that the maximum ligninolytic activities by T. versicolor were found at pH 4.0 and 5.0 [33]. Asgher et al. [40] showed that the optimum enzymes production by S. commune IBL-06 was found to be at pH 5 while pH 4.43 and 4.46 for S. commune NI-07 [41]. Change in pH will affect the three dimensional structure of laccase which in turn leads to the decrease in laccase

Production and activity of ligninolytic enzymes are also influenced by aeration. There are several purposes of aeration such as to supply oxygen into the media, for the removal of CO2, heat dissipation, distribution of water vapor to regulate humidity, and circulation of volatile compounds produced during metabolism. Thus, this factor should be optimized to improve rate of delignification [49].

Other factor such as various supplements (Cu2+, Mn2+, ferulic acid, xylidine, veratric acid, vanillic acid, cinnamic acid, guaiacol, etc.) for the SSF media have previously been reported in studying their effect on production of ligninolytic

#### Table 4.

Lignocellulosic biomass pretreatments with different white-rot fungi species and their isolated enzymes.

activities, optimal lignin degradation 29.88 0.97% (w/w) with cellulose loss 11.70 1.30% (w/w) were observed at 75% moisture content [44]. It was reported that the lignin degradation increased with increase in moisture content. Cellulose and hemicellulose degradation were found to be increased at higher moisture content and small particle size. The selectivity value, SV also influenced by the moisture content. Increase in moisture content decreased the SV, and this may be due to the decreasing of lignin degradation compared to cellulose loss caused by oxygen diffusion declining and ligninolytic enzymes inhibition. Similar condition was also reported for SSF of steam-exploded cornstalk by T. versicolor where the highest laccase activity achieved in this study was 2765.81 Ug<sup>1</sup> at 75% moisture content [33]. A study on laccase production by a novel white-rot fungus Pseudolagarobasidium acaciicola LA 1 through SSF of Parthenium biomass reported that the highest laccase activities, 16,388 Ug<sup>1</sup> of substrate was found at liquid to solid ratio of 5 with an incubation period of 7 days [42].

#### 3.4 Temperature

Temperature is another very critical factor in the pretreatment using white-rot fungi. However, different genus has different tolerant to temperature. Fungal physiology, fungal strain and types of substrate also resulted in different optimal temperature for biological pretreatment [30]. This statement is in agreement with several studies which showed the production of ligninolytic enzyme using white-rot Fungal Pretreatment of Lignocellulosic Materials DOI: http://dx.doi.org/10.5772/intechopen.84239

fungi has various optimal temperature [33, 50]. White-rot basidiomycetes grow optimally at temperature between 25 and 30°C while most of white-rot ascomycetes fungi grow optimally at 39°C [5]. The metabolism of these fungi produces heat and develops temperature gradients in SSF media. The accumulated heat can lead to adverse effect on the fungal growth and their metabolic activity which leading to the denaturation of the key enzymes. From the studies on pretreatment of rice straw, ligninolytic activities by S. commune was found to be peaked at 30 and 35°C [40, 41]. Meanwhile the highest ligninases production by T. versicolor was reported at 40°C [51]. Adekunle et al. [33] reported in their study on the SSF of steamexploded cornstalk with T. versicolor that there was a direct correlation between the temperature and laccase production, with the highest laccase activity of 2677.16 U g<sup>1</sup> was produced at 28°C. The maximum production of laccase by T. giganteum AGHP (1.53 <sup>10</sup><sup>5</sup> Ug<sup>1</sup> of dry substrate) was obtained at 30°C [39]. This study also showed that lower temperature of 10 and 20°C are not suitable for the growth of fungi due to lower enzyme production. Similar result was obtained from the study on laccase production by Pseudolagarobasidium acaciicola LA 1, the optimum production (19,944 Ug<sup>1</sup> dry substrate), was found at 30°C [42].

#### 3.5 pH

activities, optimal lignin degradation 29.88 0.97% (w/w) with cellulose loss 11.70 1.30% (w/w) were observed at 75% moisture content [44]. It was reported that the lignin degradation increased with increase in moisture content. Cellulose and hemicellulose degradation were found to be increased at higher moisture content and small particle size. The selectivity value, SV also influenced by the moisture content. Increase in moisture content decreased the SV, and this may be due to the decreasing of lignin degradation compared to cellulose loss caused by oxygen diffusion declining and ligninolytic enzymes inhibition. Similar condition was also reported for SSF of steam-exploded cornstalk by T. versicolor where the highest laccase activity achieved in this study was 2765.81 Ug<sup>1</sup> at 75% moisture content [33]. A study on laccase production by a novel white-rot fungus Pseudolagarobasidium acaciicola LA 1 through SSF of Parthenium biomass reported that the highest laccase activities, 16,388 Ug<sup>1</sup> of substrate was found at liquid to solid ratio

Lignocellulosic biomass pretreatments with different white-rot fungi species and their isolated enzymes.

Fungi species Substrate(s) Enzymes References Ceriporiopsis subvermispora Hazel branches Laccase and MnP [34]

Echinodontium taxodii Bamboo Laccase and MnP [37] Trametes versicolor Pine wood chips Laccase [14]

Tricholoma giganteum Wheat straw Laccase [39]

Pseudolagarobasidium acaciicola Parthenium biomass Laccase [42] Dichomitus squalens Chestnut shell Laccase [43] Daedalea flavida Cotton stalks Laccase and LiP [44]

> Sugarcane bagasse Sisal fiber

Stereum ostrea Wheat bran MnP [46] Pleurotus ostreatus Rice straw N.S [47] Polyporus brumalis Wheat straw N.S [48]

Corn cobs Sugarcane bagasse Wheat straw

Schizophyllum commune Banana stalk

Biomass for Bioenergy - Recent Trends and Future Challenges

Tremetes villosa Coconut shell

Albizia chips [35] Miscanthus sinensis [36]

Corn stalk [33] Corn silage [38]

Laccase, LiP and MnP [40, 41]

MnP [45]

Temperature is another very critical factor in the pretreatment using white-rot

fungi. However, different genus has different tolerant to temperature. Fungal physiology, fungal strain and types of substrate also resulted in different optimal temperature for biological pretreatment [30]. This statement is in agreement with several studies which showed the production of ligninolytic enzyme using white-rot

of 5 with an incubation period of 7 days [42].

3.4 Temperature

50

N.S,not specified.

Table 4.

pH is one of the prominent parameters in the cultivation of fungi and it is very problematic to control in SSF [52]. The initial pH of the medium influences the microbial growth and the production of ligninolytic enzyme. White-rot fungi grow well at pH 4–5, while substrate acidity decreases their growth. A study conducted on the isolation of laccase from a novel white-rot fungus Pseudolagarobasidium acaciicola LA 1 through SSF of Parthenium biomass showed that the isolated laccase was found to perform optimally at pH 4.5 and highly stable within the range of pH 4–7 for 24 h [42]. The effect of pH is important in the case of laccase production, and a small change in intracellular pH will result in a decrease of macromolecules synthesis. Patel and Gupte [39] reported that the maximal laccase production (1.27 <sup>10</sup><sup>5</sup> Ug<sup>1</sup> of dry substrate) by Tricholoma giganteum AGHP was achieved at pH 5.0. No increment in the production of enzyme was found at higher pH. This may be attributed to the poor mycelial growth at an elevated pH which may restrict the laccase production. It was reported that the maximum ligninolytic activities by T. versicolor were found at pH 4.0 and 5.0 [33]. Asgher et al. [40] showed that the optimum enzymes production by S. commune IBL-06 was found to be at pH 5 while pH 4.43 and 4.46 for S. commune NI-07 [41]. Change in pH will affect the three dimensional structure of laccase which in turn leads to the decrease in laccase activity [5].

#### 3.6 Aeration

Production and activity of ligninolytic enzymes are also influenced by aeration. There are several purposes of aeration such as to supply oxygen into the media, for the removal of CO2, heat dissipation, distribution of water vapor to regulate humidity, and circulation of volatile compounds produced during metabolism. Thus, this factor should be optimized to improve rate of delignification [49].

#### 3.7 Supplements

Other factor such as various supplements (Cu2+, Mn2+, ferulic acid, xylidine, veratric acid, vanillic acid, cinnamic acid, guaiacol, etc.) for the SSF media have previously been reported in studying their effect on production of ligninolytic

enzymes [44, 53]. Many studies reported that the copper at various concentrations influences laccase production in S. ostrea,T. pubescens, P. eryngii, and P. ostreatus [46, 54]. This is related to the role of Cu2+ that controls the transcription of laccase gene and also enhances the stability of this enzyme. Meanwhile, the concentration on Mn2+ influenced both MnP and laccase production by different Pleurotus species [55].

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pecs.2014.01.001

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