Biological pretreatment offers some conceptually important advantages such as low chemical and energy use. However, a controllable and sufficiently rapid system has not yet been found. At the same time, chemical pretreatments have also serious disadvantages in terms of the requirement for specialized corrosion resistant equipment, extensive washing, and proper disposal of chemical wastes.

Table 9.14 Selected hydrolysis and fermentation strategies

Name Description Features

Biological pretreatment is a safe and environmentally friendly method for lignin removal from lignocellulose. Biological pretreatment comprises of using micro­organisms such as brown, white, and soft-rot fungi for selective degradation of lignin and hemicellulose out of which white-rot fungi seems to be the most effective microorganism. Lignin degradation occurs through the action of lignin­degrading enzymes such as peroxidases and laccase [136]. Biological pretreat­ments are safe, environmentally friendly, and less energy intensive compared to other pretreatment methods (Table 9.15). However, the rate of hydrolytic reaction is very low and needs a great improvement to be commercially applicable. Hat — akka [68] investigated the pretreatment of wheat straw using 19 white-rot fungi and found that 35% of the wheat straw was converted to reducing sugars after 5 weeks’ pretreatment with Pleurotus ostreatus compared to only 12% conversion of the untreated straw.

Ethylene glycol solution allows to fractionation of agricultural crop residues into pulps and valuable by-products. Many no-wood materials, such as vine shoots, cotton stalks, leucaena (Leucaena leucocephala) and tagasaste (Chamaecytisus proliferus) [152, 153], palm oil tree residues [140] as well as waste newspaper [154], were subjected to ethylene glycol fractionation or pulping to obtain pulp or cellulose-rich fraction for the production of ethanol. In addition, ethylene glycol was used as modifying agents in soda [141] and kraft puilping [155], aimed at improving physical and mechanical properties of the paper sheets.

A new process has been designed to fractionation of agricultural crop residues (palm oil empty fruit bunches—EFB) for the production of pulp, lignin and he — micelluloses [140]. The obtained EFB organosolv pulp was used to produce paper, and the final properties of the resulting paper sheets were improved after refining. The black liquor showed a pH of 5.8 and a lower ash content, indicating that this liquor was easy to be treated in the subsequent stage to recover the by-products and energy. The obtained lignin with high proportion of low molecular weight lignin was claimed to be applicable as an extender or as a feedstock for the synthesis of phenol-formaldehyde resins. The solvent and by-products recovery was simulated based on 1,000 kg/h of dry raw material and solvent input flow rate

7,0 kg/h with a liquid/solid ratio of 7 (Fig. 11.4). Lignin was precipitated by adjusting pH to 2 with acidified water, and ethylene glycol was recovered by multiple distillations. By simulation with commercial software (Aspen Plus), 91% of the ethylene glycol exiting in the digester was recovered, and 88% water was obtained and recycled. In a proposed recovering scheme, lignin and sugar recoveries accounted for 22% and 35% of the original lignin and sugar in the feedstock were achieved, respectively.

Regarding the production and recovery of value-added aldehydes from lignin — containing raw materials, Fig. 12.12 shows a simplified flow sheet proposed by the research group of LSRE, working with lignin-based biorefining since the 1990s [133].The strategy is to combine reaction engineering and efficient separation processes for converting lignin from pulping spent liquors into value-added aldehydes. A portion of the by-product streams is processed to extract lignosulf — onates or lignin (acidification/precipitation, UF or LignoBoost process). The subsequent processes are based on three main steps. The first step consists on the alkaline lignin oxidation in a structured bubble column reactor as reported in Sect. 12.4.5 [120]. Then, the reactor stream follows to an ultra-filtration process leading to the separation of high molecular weight fraction of degraded lignin from

the lower molecular weight species, which goes preferentially to the permeate [135]. The permeate flows through a packed bed on acid resin in H+ form to protonate the phenolates [134]. At the end, vanillin is recovered from solution by using crystallization process.

The production of lignin-based polyurethanes elastomers and foams could be also explored. The high molecular weight fraction retained in the UF process can be considered as raw material for lignin-based polyurethanes. The production of polymers from lignin is undoubtedly an attractive approach since it can take advantage of its functional groups and macromolecular proprieties. This applica­tion has been the topic of intense research and materials with quite promising properties were already obtained [80, 83, 175].

This complete process (reaction and separation) could be integrated in a pulp and paper mill, with the possibility of diverting a fraction of liquor lignin for oxidation, producing vanillin and syringaldehyde. The unreacted lignin (after oxidation and separation of added-value chemicals) can be reintroduced in the liquor stream to be burned, recovering by this way part of the energy lost by the deviated fraction. Alternatively, this lignin could be the raw material for polymers production [174]. Moreover, this process perfectly fits into the scope of new emerging lignocellulosic-based biorefineries to valorize lignin. This concept is entirely related to the development strategies and policies regulated by Agenda 21 program, offering a framework to enable the smooth transition toward a Bio-based Economy supported by innovation and sustainable growth.

Acknowledgments Authors are grateful to Dr. Detlef Schmiedl, Fraunhofer Institute for Chemical Technology, Germany and Dr. Daniel Araujo, Faculty of Engineering, University of Porto, Portugal, for kindly providing figures and data.

pH is the one of the most important factors that should be optimized for dark fermentative hydrogen production. It can affect the hydrogen production yields and also by-product formations. For dark fermentative hydrogen production optimum pH values are said to be between 5 and 6. It is important to stabilize the pH in the system because the acetic and butyric acids produced during hydrogen production will lower the pH to inhibitory acidic values. Since acid and base treatments are the kinds of pre-treatment operations pH changes in the system can change the microbial community too. Before starting continuous or large-scale applications it would be useful to determine the best pH value of the system in small-scale batch experiments [88].

Temperature can affect both microbial community and hydrogen production. Many of the studies on dark fermentative hydrogen production were conducted at mesophilic temperatures but some studies have shown that hydrogen production rates can be higher at thermophilic conditions. But from the view of economical costs thermophilic operation may not be an economically viable solution.

12.2.3.1 Origin and Isolation

Organosolv lignins are those derived from delignification processes using an organic solvent, frequently ethanol or methanol, and an acid catalyst (mineral or organic), leading to liberation of lignin from cellulosic fibers. High temperatures (approximately 195°C) and pressures (about 28 bars) lead to the cleavage of a- and b-ether linkages of lignin structure [66] and some linkages between lignin and other cell wall components. As for other pulping processes, hardwoods are more readily delignified than softwoods. At lab scale, the isolation is usually performed by acidification of resulting lignin solution and precipitation with water. The solids are recovered by centrifugation or filtration and dried [67]. At industrial scale, the liquor lignin is recovered by precipitation with an aqueous process stream, fol­lowed by filtration, washing and drying [68]. The yields reported are considerably high [67, 69, 70].

They are fast growing plants that can be easily cultivated using established agronomic practices which compensate for their relatively low capacity of metal accumulation. Their metal uptake capacity can further be enhanced by adding

conditioning fluid containing a chelator or another agent to soil to upsurge metal solubility or mobilization so that the plants can absorb them more easily. This is known as chemically induced/assisted phytoextraction. Afterwards, the soluble metal (desorbed from soil particles) is easily transported to roots surface via diffusion and translocated from roots to shoots. Complexing with organic ligands, which may occur at any point along the transport pathway, converts the metal into less toxic form thus conferring high metal tolerance in biomass plants [5]. A wide range of synthetic chelates [e. g.Ethylenediaminetetraacetic acid (EDTA), 1,2-cyclohexylenedinitrilotetraacetic acid (CDTA), diethylenetriamine- pentaacetic acid (DTPA), EGTA, EDHA, hydroxyethylethylenediaminetriacetic acid (HEDTA), nitriloacetic acid (NTA), and organic acids (e. g. citric acid, oxalic acid, malic acid) are used for enhancing root uptake and translocation of metal contaminants from soil to biomass plants, thereby improving phytoex­traction [6] (Fig. 14.3, 14.4, 14.5).

Fig. 14.4 Willows

Fig. 14.5 Poplars

Example: Indian mustard, sunflower, and maize as high biomass crop plants, willows, and poplars as high biomass trees

Advantages. The main advantage of phytoextraction is environmental friend­liness. Traditional methods that are used for cleaning up heavy metal contaminated soil disrupt the soil structure and reduce its productivity, whereas phytoextraction can clean up the soil without causing any kind of harm to soil quality. Add on benefit of phytoextraction is that it is less expensive than any other clean-up process (Fig. 14.6) (Table 14.1).

Fig. 14.6 Phytoextraction

Table 14.1 Main characteristics of the two categories of plants for phytoextraction of metals [4]: Chemically assisted phytoextraction Natural phytoextraction

Ethanol fractionation can be operated under low and medium severity as a pre­treatment process to obtain hydrolyzable cellulose. In this case, the hydrolysis reaction mainly occurred at carbon position of the side chains of lignin. Cleavage of a-aryl ether is a main reaction, which lead to the formation of a benzylic carbocation in acidic medium. The benzylic carbocation can react with water or ethanol, or form a bond with an electron-rich carbon atom in the aromatic ring of another lignin unit [27]. This reaction mechanism is supported by lignin model

compound study, in which a-aryl ether linkages are more easily degraded than b-aryl ether linkages [28]. Under highly serious conditions, b-aryl ether linkages are extensively cleaved, which is the controlling reaction in delignification. The extensive cleavage of b-aryl ether linkages results in a substantial increase of phenolic hydroxyl groups, which is confirmed by the low intensity of Cb and Cy signals in the dissolved lignin as compared to MWL [27]. After acidolysis of the ethanol dissolved lignin fraction, the contents of phenolic hydroxyl groups increased significantly, suggesting the presence of intact b-O-4 bonds in the dis­solved lignin [29]. The presence of b-O-4 structures in ethanol lignin was also demonstrated by HMQC 2D NMR [30]. In addition, the presence of carbonyl groups in the dissolved lignin indicated that the formation of Hibbert’s ketones during the fractionation process [31].

During the cleavage of b-O-4 bonds, the homolytic cleavage occurs via methide intermediate thus causes the formation of b-1 inter-linkage through radical cou­pling, which then in turn degrades under the acidic medium to give stilbenes through the loss of the y-methylol group of formaldehyde [28, 32]. In addition, b-5 units are also converted into stilbenes through the same degradation pathway [33]. With respect to cinnamyl alcohol, it is converted into ethyl ether structure [33]. In a recent report, a marked decrease of aliphatic OH and a significant increase of phenolic OH are found in ethanol dissolved lignin of Miscanthus with increase of the severity of the treatment [27]. This observation can be attributed to two simultaneous and opposite reactions: the production of p-hydroxyphenyl OH group due to the scission of b-O-4 bonds involving H units and hydrolysis of a fraction of p-coumaryl ester residues [34].

With respect to the activation energies for cleavages of the two major linkages in lignin, the study of the lignin model compounds indicates that the activation energies for cleavages of a-aryl ethers bonds range from 80 to 118kJ/mol, depending on substituent [35]. These values are slightly higher than those in both auto-catalyzed and acid-catalyzed acetic acid fractionation processes, which are

78.8 and 69.7 kJ/mol, respectively [36]. However, the reported activation energy for b-aryl ether hydrolysis is 150 kJ/mol [27]. Obviously, the high value was not considered to be the controlling reaction in the ethanol fractionation process.

Lignin condensation is an important counterproductive reaction in an acidic or alkaline ethanol fractionation process. The intermediates, i. e., reactive benzyl carbocations or benzyl-linked oxygen atoms, can form a bond with an electron — rich carbon atom in the aromatic ring of another lignin units resulting in the production of condensed products. It has been reported that in a weak acid system, protonation of a benzyl-linked O atom was a SN2 type reaction [37].

The effect of lignin concentration on the reaction rate of vanillin production is depicted in Fig. 12.9c. The calculated maximum yield on lignin basis (wt%) decreases with the CL: 10, 8.3, and 3.0% for CL of 30, 60, and 120 g/l, respec­tively. The slope of the straight line found for initial vanillin production rate as a function of the initial lignin concentration provides the reaction order of 1 with respect to lignin concentration [116].

Considering the results of vanillin production from a kraft lignin in alkaline medium, under the reported conditions, the following kinetic law was achieved:

rv = k[O2]175 [L] (12.1)

Biomining of copper. Copper was the first metal extracted by biomining. During the period 1950-1980, as compared to conventional metallurgical techniques, biomining appeared as economically viable and potential technology to recover Cu

Fig. 14.15 Metal specific chelating resin

from low grade ore, like copper sulfide. It has been reported that the Lo Aguirre mine in Chile processed about 16,000 t ore per day between 1980 and 1996 using biomining [27].

Fungal leaching of manganese ore. Recovery of Mn from low grade ore of Mn by using pyrometallurgical and hydrometallurgical methods is expensive because of high energy and capital inputs. Besides, it also contributes a lot to environ­mental pollution. On the other hand biomining of Mn from manganiferous ores using microbial leaching is cost effective as well as environment friendly. It has
been reported that a fungus Penicillium citrium can solubilize or extract 64.6% of Mn from the low grade ore [28].

Biomining of gold. Using cyanide method, it is very much difficult to extract gold, when gold is covered with insoluble metal sulfides. Biomining of these sulfide films is the best option to achieve satisfactory gold recovery. Gold extraction plants of Sao Benzo in Brazil, Ashanti in Ghana, Tamboraque in Peru are known to have such biomining facilities. A series of demonstration plants was also commissioned during 2002 in the Hutti Gold Mines in Karnataka [27].

Recovery of chromium from tannery sludge. About 40% of total Cr used in tanning industry end up in the sludge. Cr is non-biodegradable and can easily accumulate in food chain causing serious health effects to human beings. Use of microfungi due to their biochemistry and relatively high immunity to hostile conditions such as pH, temperature etc. provide a better alternative to commercial leaching processes. It has been demonstrated that chromium from tannery sludge can be bioleached up to 99.7% using indigenous acidophilic fungi, A. thiooxidans [29]. Another Cr recovery option from tannery waste is to grow potential Cr accumulating fungi in tannery waste and subsequent extraction of Cr from the harvested biomass. In an extensive study on Cr accumulation by fungal biomass, the author identified a fungal strain, Paeciomyces lilacinus which can accumulate Cr up to 18.9% of their dry biomass [30].

Bioleaching of economical metals from electronic and galvanic waste. These contain various valuable metals. Microbial process involving both bacteria and fungi, which produce inorganic and inorganic acids, can mobilize these metals from the waste. Metals such as Al, Ni, Pb, and Zn have been reported to be extracted by this process. Microbial leaching has also been found effective to recover Ni and Cd from spent batteries [31].

Phytoextraction of metal. Phytoextraction of metals from low or moderately contaminated soil or waste material is recommended but not an option for highly contaminated soil. In later case, it may take decades or even centuries to reduce the contaminant concentration to an acceptable limit. Instead of using low biomass hyperaccumulator plants, high yielding plants along with addition of chelating agent proved to be better method to phytoextract metal from soil. Uses of different plants in chelant-induced phytoextractiopn are summarized in Table 14.3.

However, often application of chelants can result in residual toxicity in soil on which it is applied. Thus, natural accumulation of metals would be the best option provided application of mycorrhizal fungi, plant growth promoting rhizobacteria and other beneficial microbes in soil that can enhance the efficiency of extraction processes [32]. It has also been reported that plants colonized by the AM fungi not only enhance growth, but also significantly increase Pb uptake in root and higher translocation to the shoot at all given treatments [33]. It has also been seen that three mycorriza inoculated plant glomus species namely G. lamellosum, G. intraradices, G. proliferum and their consortia greatly enhance accumulation of Cr from tannery waste to plants.

Thermal pretreatment for fractionation and solubilization studies of lignocellulosic materials have shown the efficiency to improve the yields of extraction of hemi — celluloses. Boussarsar [20] have evaluated the SCB conversion by hydrothermal treatment. Optimal conditions were 170°C for 2 h, reaching higher solubilization of hemicellulose than that at 150°C and lower degradation of sugar monomers than at 190°C. However, analysis of thermal hydrolysates shows the presence of xylan oligomers and polymers with large chains. On the other hand, Sendelius [166] has evaluated the steam pretreatment conditions with respect to final ethanol yield, using SCB as feedstock. The variables considered were temperature (180, 190, and 205°C), time (5 and 10 min), and impregnating agents (water, 2% SO2 by weight of water in the bagasse and 0.25 g H2SO4 per 100 g dry matter). The most prominent tested pretreatment condition was: SO2-impregnation at a temperature of 180°C during 5 min, which gave a glucose yields in average 86.3% and xylose yields in average 72.0%. The fermentation of these hydrolyzed materials gave an overall ethanol yield of 80%, based on theoretical value.

Wet Oxidation

Wet oxidation (WO) is the process of treating material with water and either air or oxygen at temperatures above 120°C. Two types of reactions occur during WO: a low temperature hydrolytic reaction and a high temperature oxidative reaction. It has been demonstrated that a combination of alkali and WO reduces the formation of toxic furfuraldehydes and phenol aldehydes [97]. Martin [118] have investi­gated different conditions pH, temperature, and reaction time of WO pretreatment on fractionation and enzymatic convertibility of SCB, while pressure (12 bar) was kept constant. The highest cellulose content, nearly 70%, was obtained in the pretreatment at 195°C, 15 min and alkaline pH. The highest sugar yield in the liquid fraction, 16.1 g/100 g, was obtained at 185°C; 5 min and acidic pH. Although the analysis of the solid fraction in most of the pretreatments showed high degrees of hemicelluloses solubilization, the content of free sugars in the liquid fraction was very low. It is known that WO mainly catalyzes the transfer of hemicelluloses from the solid phase to the liquid phase, but it does not catalyze the hydrolysis of the liberated hemicelluloses molecules. The products of hemi — celluloses hydrolysis during WO are not monosaccharides, but sugar oligomers.