da Costa Lopes et al. Sustainable Chemical Processes 2013, 1:3http://www.sustainablechemicalprocesses.com/content/1/3/3
REVIEW Open Access
Ionic liquids as a tool for lignocellulosic biomassfractionationAndre M da Costa Lopes1, Karen G João1, Ana Rita C Morais1, Ewa Bogel-Łukasik2 and Rafał Bogel-Łukasik1*
Abstract
Lignocellulosic biomass composes a diversity of feedstock raw materials representing an abundant and renewablecarbon source. In majority lignocellulose is constituted by carbohydrate macromolecules, namely cellulose andhemicellulose, and by lignin, a polyphenilpropanoid macromolecule. Between these biomacromolecules, there areseveral covalent and non-covalent interactions defining an intricate, complex and rigid structure of lignocellulose.The deconstruction of the lignocellulosic biomass makes these fractions susceptible for easier transformation tolarge number of commodities including energy, chemicals and material within the concept of biorefinery. Generally,the biomass pre-treatment depends on the final goal in the biomass processing. The recalcitrance of lignocellulosematerials is the main limitation of its processing once the inherent costs are excessively high for the conventionalpre-treatments. Furthermore, none of the currently known processes is highly selective and efficient for thesatisfactory and versatile use, thus, new methodologies are still studied broadly. The ionic liquid technology onbiomass processing is relatively recent and first studies were focused on the lignocellulosic biomass dissolution indifferent ionic liquids (ILs). The dissolution in IL drives to the structural changes in the regenerated biomass byreduction of cellulose crystallinity and lignin content contrasting to the original biomass. These findings providedILs as tools to perform biomass pre-treatment and the advantageous use of their specific properties over theconventional pre-treatment processes. This review shows the critical outlook on the study of biomass dissolutionand changes occurred in the biomass during this process as well as on the influence of several crucial parametersthat govern the dissolution and further pre-treatment process. The review of currently known methods of biomassfractionation in IL and aqueous-IL mixtures is also discussed here and perspectives regarding these topics aregiven as well.
IntroductionLignocelluloses such as hardwood, softwood and agricul-ture residues are the main raw material from renewablefeedstock representing an abundant carbon source [1].The main components of lignocellulosic materials arecellulose, hemicellulose and lignin. Cellulose and hemi-cellulose constitute the carbohydrate fraction wherecellulose is a linear semi-crystalline fibrous homopoly-saccharide and hemicellulose is a branched amorphousheteropolysaccharide. Cellulose chains are embedded inthe core of the lignocellulosic biomass with hydroxylgroups oriented to form strong intra- and intermolecular
* Correspondence: [email protected]ório Nacional de Energia e Geologia, Unidade de Bioenergia,1649-038 Lisbon, PortugalFull list of author information is available at the end of the article
hydrogen bonds. This lattice network results in orga-nised micro- and macrofibrils as well as interacts withthe hemicellulose fraction. Lignin is characterised as anamorphous phenylpropanoid polymer that acts as a“glue” binding hemicellulose and cellulose by ester link-ages and hydrogen bonds, respectively [2]. These mo-lecular interactions define a complex and rigid structureforming a recalcitrant material against hydrolysis guidingto difficult, harsh and expensive separation processes [3].Generally, the separation process of these compounds
is performed by thermochemical and/or physical me-thods as a pre-treatment, exposing the carbohydratefraction to facilitate further processing. Carbohydratesare subsequently hydrolysed to sugar monomers andthen converted to bioalcohols (ethanol, butanol), car-boxylic acids, biohydrogen or methane by fermentation
try Central Ltd. This is an Open Access article distributed under the terms of the/creativecommons.org/licenses/by/2.0), which permits unrestricted use,, provided the original work is properly cited.
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processes [4,5]. On the other hand, the extracted lignincan be used as a low-value heating fuel, binder, dispers-ant, emulsifier and sequestrant. Lignin can also act as asource of high-value molecules, such as biologically ac-tive polyphenols and tannins [6,7]. The conventionalmethodologies have technological limitations such as aninsufficient selectivity and partial degeneration of prod-ucts that compromise the efficiency of the separationprocesses. Hence, the current and envisaged investi-gations are focused on understanding the pathways toimprove the selective separation of lignocellulose com-pounds to accomplish feasible and sustainable processes.Ionic liquids (ILs) are recognised to facilitate more
green applications in reactions and separations due totheir unique beneficial properties, such as negligiblevapour pressure [8,9] and high thermal stability [10].Their very low vapour pressure reduces the risk ofexposure that is a clear advantage over the use of theclassical volatile solvents. Although, studies of basicproperties in the environmental risk assessment of ILsare still required [11], it is notable that toxicity of theILs is mainly ascribed to the alkyl chain and that thetoxicity of imidazolium and pyridinium ILs increaseswith their cation chain length [12,13]. ILs are com-pounds composed solely of ions with immeasurablecombinations of anions and cations. They possess widelytuneable properties, such as hydrophobicity [14], polarityand solvent power [15,16].In the last decade, numerous studies focused on the
dissolution of natural polymers in ILs demonstrating agreat potential of ILs as solvents [17-21]. Cellulose wasone of the most studied biopolymers exhibiting a highsolubility in a variety of ILs [22]. A wide range of carbo-hydrate solubilities was scrutinised using different ILs,presenting that one of the main benefits of using ILs todissolve carbohydrates is that ILs can be tailored to ac-complish dissolution or functionalisation of these poly-mers [22].Due to the ILs solvent power, their use in the develop-
ment of alternative methods for the extraction and pro-cessing of carbohydrates and other compounds fromlignocellulosic biomass was recently explored intensively.The aim of this work is to describe the current state-of-the-art on the fractionation of lignocellulosic biomass inILs as a pre-treatment methodology.The cations and anions of ILs used in this review are
presented in Figures 1 and 2, respectively.
Biomass dissolutionInitially, Fort et al. demonstrated that solvent systemsbased on [bmim][Cl]/DMSO-d6 (84/16 wt%) are capableto partially dissolve wood chips [23]. They describedswelling and the size reduction of wood particles du-ring the dissolution basing on the colour intensity and
viscosity of the solution mixture. Meanwhile, Kilpelainenet al. reported a complete dissolution of 8 wt% of driedwood sawdust samples (Norway spruce and Southernpine) in both [bmim][Cl] and [amim][Cl] ILs, in thetemperature range from 80 to 130°C after 8 hours [24].The complete dissolution of wood in these ILs was at-tributed to the water content and the particle size ofwood samples. Water was found to significantly reducethe solubility of wood in ILs [19] and smaller particleswere reported as better to be dissolved in water [24].Furthermore, Sun et al. demonstrated that both soft-wood (southern yellow pine) and hardwood (red oak)can be dissolved in [emim][CH3COO] [25]. More than90% (w/w) of the added wood was dissolved after mildgrinding, at 110°C within 16 hours. The authors con-cluded that [emim][CH3COO] is more efficient in dis-solving biomass than [bmim][Cl] and [amim][Cl]. Ahigh-throughput screening in the dissolution of celluloseand wood chips in ILs with 5% (w/w) at 90°C for12 hours was made by Spiess et al. [26]. The investiga-tion was performed at identical conditions in a singleexperiment to allow a comparison of a variety of ILs.They found out that [emim][Cl], [bmim][Cl] and [emim][Et2PO4] were only able to partially dissolve differentwood chips (spruce, silver fir, common beech and chest-nut). Additionally, [emim][CH3COO] was defined as themost efficient for dissolving cellulose, and [amim][Cl]was found to be the most suitable for use in biomass,dissolving all tested wood. Only Silver fir was not com-pletely dissolved in [emim][CH3COO] [26]. By heatingwheat straw and pine wood at 100°C after 24 hours,[emim][CH3COO] demonstrated to be a poor solvent[27]. At the same conditions, [bmim][Cl] was the mostefficient in the dissolution of straw and [emim][Cl]tended to be the most promising solvent for both typesof biomass. For the best results, the additional investiga-tion with transparent solutions did not exclude presenceof biomass fibres after filtration indicating an incompletedissolution [27]. In fact, Lee et al. stated a very lowsolubility of wood flour (< 5 g/kg) and a high solubilityof microcrystalline cellulose (> 100 g/kg) in [emim][CH3COO], proving that the presence of lignin restrictsthe solubility of wood flour in this IL [28]. In the dis-solution of beech and spruce wood at 115°C, [emim][CH3COO] showed to be more suitable for the first one,but complete dissolution was observed for none of theexamined biomasses even during a 72-hour process [29].An incomplete dissolution of pine wood in this IL at120°C and 155°C within 3 hours was also reported [30].These findings were the opposite of those presentedearlier by Singh et al [31]. Basing on the microscopic ob-servations, Torr et al. clearly demonstrated that [emim][CH3COO] is capable to completely dissolve all majorcomponents found in the plant cells of switchgrass after
Figure 1 Cations referred in this work listed by order of appearance. [bmim] - 1-butyl-3-methylimidazolium, [emim] - 1-ethyl-3-methylimidazolium, [amim] - 1-allyl-3-methylimidazolium, [Ch] - cholinium, [bzmim] - 1-benzyl-3-methylimidazolium, [HEA] - 2-hydroxyethylammonium, [bmpy] - 1-butyl-3-methylpyridinium, [Me(OEt)3Et3N] - triethyl-(2-(2-methoxyethoxy)ethoxy)ethylammonium,[DMEA] - N,N-dimethylethanolammonium, [mmim] - 1,3-dimethylimidazolium, [hmim] - 1-hexyl-3-methylimidazolium, [pmim] - 1-propyl-3-methylimidazolium, [abim] - 1-allyl-3-butylimidazolium, [eMeOHpy] - 1-ethyl-3-(hydroxymethyl)pyridine, [bHim] - 1-butylimidazolium.
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3 hours at 120°C [31]. On the other hand, [bmim][CH3COO] was tested to dissolve poplar wood and adissolution yield of 96 wt% was obtained at 130°C after12 hours, while [bmim][Cl] only achieved 23 wt% ofpoplar dissolution [32]. This indicates that [CH3COO]anion has a great influence on the dissolution of bio-mass. In the dissolution of sugarcane bagasse at 100°Cwithin 0.5-8 hours, once again [emim][CH3COO] dem-onstrated to be more effective than other ILs such as[bmim][Cl] and [emim][DEP] [33].A reduced time of biomass dissolution was attained
using microwave or ultrasound irradiation instead ofconventional heating [25,34]. The enhancement in thedissolution obtained was attributed to the ionic structureof ILs that absorbs efficiently these types of irradiation.In consequence the frequency of collisions between the
anions and cations of ILs and the wood macromoleculesincreases leading to fast and effective dissolutions [34].Application of a prior ball-milling treatment to thebiomass in order to reduce the molecular weight ofthe lignocellulosic complex can also improve dissolution[35,36]. Unfortunately, these mechanical treatment pro-cesses are low effective and are inappropriate for an in-dustrial application.To understand the rules governing the fractiona-
tion process, the use of polarity approach can be consi-dered. To do so, the quantitative Kamlet-Taft parameters[37-39] were broadly used in the literature. Using Kamlet-Taft parameters it was found that the anion basicity influ-ences the lignocellulosic biomass dissolution [40,41]. Acorrelation between higher dissolution capacities of ILsand higher value of β parameter was determined [40].
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Brandt et al. observed high dissolution power for [bmim][CH3COO], [bmim][Cl] and [bmim][DMP] ILs. Theydemonstrated that acetate IL is the most effective forswelling and dissolution [40]. This is justified by a veryhigh β value exhibited by [CH3COO] anion (1.20), whichis a considerably stronger hydrogen bond acceptor. On
the contrary, [bmim][OTf] with a low β value (0.48) re-vealed a very poor swelling process, starting at 120°C. Theswelling was also slow with [bmim][MeSO4] and [bmim][Me2PO4]. Even worst dissolution was demonstrated byusing [bmim][N(CN)2] that dissolved neither cellulosenor lignocellulosic material [40]. Other, [emim][TFA] and
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[Ch][PrOO] ILs showed no evidence of dissolving ligno-cellulose as they were tested to dissolve bamboo biomasswithout success even after 24 hours at 120°C [42]. On theother hand, [emim][Gly], an amino-acid based IL had theability to dissolve the bamboo biomass due to a highhydrogen bond basicity of amino-acid derived anions. Af-ter 8 hours at the same temperature, complete dissolutionof bamboo with [emim][Gly] was observed [42].The presented results are affected by different vari-
ables of the IL dissolution process including the typeand size of biomass, IL/biomass ratio, temperature andtime of dissolution, water content in the solution mix-ture and others. The complete biomass dissolution inILs is very dependent upon these factors which, in turn,should be optimised for the specific IL-biomass dis-solution process. Nevertheless, [bmim][Cl], [amim][Cl],[emim][Cl] and [emim][CH3COO] demonstrated to bethe most used and efficient ILs in dissolving lignocellu-losic biomass, considerably. The [emim][CH3COO] isgenerally recommended since it exhibits minor environ-mental issues comparing to ILs composed by halideanion [43,44]. The biomass dissolution in ILs has beenreviewed [45,46] and there is still room for progress inthe dissolution performance by seeking new ILs with im-proved properties.
The influence of ILs’ properties on the biomass dissolutionThe ability of ILs to dissolve carbohydrates and lignin isconsidered as an effective disruption of the intricatenetwork of non-covalent interactions between thesepolymers. Rogers and co-workers suggested that a highchloride concentration and activity of [bmim][Cl] is re-sponsible for breaking an extensive and well organisedhydrogen-bonding network of cellulose and thus pro-moting dissolution [19]. Remsing et al. using 13C and35/37Cl NMR demonstrated that the interaction betweenthe carbohydrate and the anion of an IL is predominantcompared to the interactions of carbohydrate with thecation [47]. It was reported that the dissolution of carbo-hydrates in [bmim][Cl] involves the formation of hydro-gen bonds between chloride ions of the IL and hydroxylprotons of sugar units from carbohydrates in a 1:1 stoi-chiometry. The IL cation has also some influence on thedissolution by interacting with cellulose hydroxyl oxygengroups [34]. In the case of lignocellulose dissolution theprinciple is the same, once the main fraction of thesematerials comprises carbohydrates [31]. However, thepresence of lignin and extractives in lignocellulose re-stricts the solubility and the appearance of a brownishviscous mixture solution is observed during the process[26]. ILs not only disrupt the hydrogen bonding inter-action of crystalline cellulose in wood, but also interactand solvate the aromatic characters of lignin by π-π andn-π interactions, generally performed by the IL cation
[34,48,49]. This phenomenon is enhanced by a more ef-fective stirring that increases the hydrodynamic stress ofthe lignocellulosic particles and mass transfer to theparticle surface improving the dissolution process [26].The microwave irradiation also enhances the dissolution,since depolymerisation of cross-linked phenylpropanoidunits of lignin and hydrogen bonding interactions be-tween the wood matrix and the IL anion are intensified[34]. The ultrasound irradiation seems to induce thesame effect on biomass dissolution [50]. On the otherhand, by breaking the native structure with ball-milling,a reduction of molecular weights of lignin and carbohy-drate polymers is observed [35]. Thus, an effective re-duction of polymers leads to a subsequent dissolutionenhancement in ILs.The selection of ILs for lignocellulosic biomass dissol-
ution is difficult, due to the different physical and chem-ical properties that IL presents. It was referred that ILsconstructed by bulky cation and halide anion may de-crease the concentration of active chloride ion and thusthe solvating capacity for both cellulose and lignin is re-duced [19]. The comparison of the efficiency of [emim][Cl] and [bmim][Cl] in the dissolution of rice straw al-lows to notice that [emim] cation, as smaller sized than[bmim] cation, might be more effective due to its higherdegree of interaction with the cellulose chain [33,51]. Asmaller anion is also preferable to be able to diffusefaster within the lignocellulosic matrix as in the case ofchloride anion. Nevertheless, the improvements in ligno-cellulose dissolution are related with the hydrogen bondbasicity of the IL anion as referred above. ILs with astrong hydrogen bond basicity are effective in weakeningthe hydrogen-bonding network of the polymer chains[40,52]. As it was stated before the increased basicity ofthe [CH3COO] anion makes it more efficient at dis-rupting the inter- and intramolecular hydrogen bondingin biopolymers than Cl anion [25]. To predict the pre-treatment efficiency of a certain IL in disrupting ligno-cellulose, β Kamlet-Taft parameter is an useful tool asdemonstrated by Brandt et al. for [bmim][CH3COO]and [bmim][Cl] [40]. As it was aforementioned, theyconfirmed better efficiency of [CH3COO] IL (β =1.20)than the chloride IL (β =0.83) in the dissolution of pinewood chips. Indeed, ILs with β ≥ 1.0 support high yieldsof fermentable sugars following pre-treatment [41]. As itis stated above, glycinate ionic liquid demonstrates highhydrogen basicity. The glycinate anion is characterisedto have a weak electron withdrawing group (−NH2) on βcarbon, although this negative inductive effect does notinfluence the hydrogen bond formation by the carboxyl-ate group (−COO-) with lignocellulosic biomass [42]. Inthe case of [emim][TFA] the strong electron withdraw-ing groups (−F-) decrease the electron density of COO-,thereby it reduces the formation of hydrogen bonds
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between biomass and IL during dissolution [42]. ILs withanions showing a low hydrogen bond basicity, such as[bmim][MeSO4], [bmim][HSO4], [bmim][MeSO3] areunable to perform a complete dissolution of the bio-mass, since interaction with cellulose is weak [53]. Otherexamples are [bmim][OTf] and [bmim][N(CN)2] whichare incapable to break the hydrogen bonds in the matrixand therefore do not expand it at all or only minimally[40]. Generally, increased hydrogen bond basicity of theanion leads to the incorporation of water molecules inthe IL structure [54], reducing the dissolution of bio-mass in IL. Therefore, drying of the IL prior to use is re-quired regardless the biomass moisture. However, notonly hydrogen basicity of IL is important, but also itsstructure affects the dissolution process. Muhammadet al. presented that the [Ch][PrOO] IL, in spite of ahigh hydrogen basicity, has a poor dissolution ability[42]. This behaviour was explained by the internal inter-action between the hydroxyl end group of IL cation andits anion. Furthermore, the absence of aromatic ring inthe cation structure had a negative effect on the dissol-ution unable to interact with lignin.The viscosity of ILs is also an important parameter, be-
cause it can impact the mixing and mass transfer oflignocellulose and IL itself. Abe et al. verified that ILscharacterised by low viscosity are able to extract higheramounts of carbohydrates from the bran [52]. This mayoccur due to better dispersion state of bran in less vis-cous ILs. It was also reported that ILs with an adequatepolarity and a low viscosity demonstrate good ability toextract polysaccharides in a short time. They showedthat the low viscosity and highly polar IL, [emim][PO(O)H2], allows for rapid extraction of cellulose and othercarbohydrates from bran under mild conditions [52].Kilpelainen et al. attempted the dissolution of wood in[benzylmim][Cl], once the benzyl ring from the IL cationcould interact with the phenyl aromatic rings of ligninby π-π interactions [24]. In fact, a major drawback of[benzylmim][Cl] application is related to high viscosityof this IL [24].The lower melting point of [emim][CH3COO], in
comparison with [emim][Cl] and [bmim][Cl], also facili-tate the dissolution of biomass and handling of the mix-ture, which makes [emim][CH3COO] a better solventthan chloride-based ILs in biomass processing [25]. It isworth noting that, at room temperature, [emim][Cl] and[bmim][Cl] are in solid state in contrast to the liquidstate of [emim][CH3COO]. Therefore, higher tempera-tures are required for [emim][Cl] and [bmim][Cl], al-though even then these ILs present high viscosities [19].
The biomass regeneration processDissolution step is the initial step guaranteeing thesuccessful processing of biomass. However, due to the
complex matrix of biomass, the adequate recovery ofeach fraction is needed for the production of energy,chemicals and biomaterials in the frame of biorefineryconcept. Generally, the process guiding to the biomassfractions’ recovery occurs as follows: at initial stage, lig-nocellulosic biomass is generally added to an IL, with acertain solid/liquid ratio, and is treated at the specifiedtemperature and time applying a mechanical stirring tothe mixture. From the literature review presented above,the dissolution can be partial or complete, although it isrecommended to drive to the complete biomass dissol-ution to improve the pre-treatment efficiency. The nextstep is one of the most important as it corresponds tothe regeneration of biomass fractions with the ad-dition of a precipitating solvent (also designated asantisolvent), such as water, acetone, dichloromethane,and acetonitrile [23]. Fort et al. described the disso-lution of wood chips in [bmim][Cl] with addition ofthe referred antisolvents [23]. They obtained the rege-nerated biomass fraction composed essentially by cel-lulose. The regenerated cellulose-rich material wasfound to have purities, physical properties, and pro-cessing characteristics comparable to those of cel-lulose standard. The regeneration yield of biomassranged from 30 to 60 wt%, for a pre-treatment tem-perature of 100°C during 16 hours [23]. The regene-ration yields presented in all the reviewed works arestrongly dependent on the IL, biomass, antisolvent andpre-treatment conditions used. The regeneration yieldsobtained are depicted in Tables 1, 2 and Additional file 1:Table S1. Wang et al. stated that the addition of waterresulted in the reconstitution of a brown gel [34]. Aftergel filtration a clear and dark brown solution was obtainedand by washing with water/DMSO several times, the gelbecame white indicating the main presence of carbohy-drates [34].After the regeneration, the crystallinity of cellulose is
abruptly reduced in comparison with that of cellulosefrom the original lignocellulosic material. Cellulose inthe pre-treated biomass is no longer constrained in arigid structure and has similar level of the molecularflexibility as the surrounding hemicellulose [30]. By mi-croscopic observations, Singh et al. verified the precipi-tation of fibrous structures of cellulose (300–500 mMlong) after addition of water [31]. To assist the lignin re-moval from the regenerated product, fluorescence tech-niques were used. Disruption of the lignin–carbohydratecomplex occurred simultaneously with cellulose crystal-linity reduction [31]. Analysing X-Ray diffraction (XRD)patterns of the regenerated material from bamboo pre-treatment, Muhammad et al. observed a clear change incrystallinity of cellulose in dissolution and regenerationprocesses [42]. Li et al. compared the pre-treatment ofthe switchgrass using [emim][CH3COO] and a dilute
C – cellulose; H – hemicellulose; L – lignin; O – others; a dried in vacuum for 8 h; b dried at 105°C, for 12 h; c dried under vacuum at 40°C for 48 h; d glucose yield after enzymatic hydrolysis; e 16 wt% DMSO; f used asreceived; g dried at 105°C overnight; h under microwave irradiation; i glucan basis; j xylan, mannan, galactan and arabinan basis; k ash basis; l dried overnight 90°C.
C – cellulose; H – hemicellulose; L – lignin; O – others; a dried under vacuum at 40°C for 48 h; b glucose yield after enzymatic hydrolysis; c 16 wt% DMSO; d used as received; e dried at 105°C overnight; f glucose yieldafter enzymatic hydrolysis regarding the original glucan content; g xylose yield after enzymatic hydrolysis regarding the original xylan content; h vacuum dried at room temperature for 12 h; i dried at 100°C for 12 h;j glucan basis; k xylan, mannan, galactan and arabinan basis; l ash basis; m dried overnight 90°C; n xylan basis; o reducing sugar yield regarding the initial content in pre-treated biomass; p dried overnight in a vacuumoven at 100°C; q lignin yield considering the total biomass input.
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acid [62]. For both untreated and dilute acid pre-treatedswitchgrass samples little or no change in cellulose crys-tallinity was observed, but for the sample obtained afterpre-treatment with IL the crystallinity was altered signifi-cantly. They concluded that cellulose could suffer a struc-tural transformation from cellulose I to cellulose II thatprovides higher performances of enzymatic hydrolysis. Ifthe IL is not efficient in breaking the interactions betweenlignin and other wood components from cellulose, thestructural transformation of cellulose I to cellulose II isnot be observed [62]. The crystallinity index (CrI) calcu-lated by XRD for the IL pre-treatment switchgrass wasfound to be significantly lower (2.6) than that for bothuntreated (26.2) and dilute acid pre-treated (39.1) switch-grass. Contrary to Singh et al. observations, the IL pre-treated switchgrass showed no fibrous structure, but aporous surface was observed over several length scales, byscanning electron microscope [62]. In fact, a less crystal-line form of pre-treated samples with [emim][CH3COO]is typical, since similar changes were observed for differentbiomass, namely corn stover, maple wood flour, poplarand switchgrass using the same IL [41]. The [emim][CH3COO] demonstrated to be more effective than[emim][Cl], [amim][Cl] [bmim][Cl] and [HEA][HCOO] inreducing the crystallinity of cotton stalks for different par-ticle sizes tested [63,64]. The [emim][Cl] was verified toneed more time to effectively disrupt the crystalline struc-ture of biomass in comparison with [emim][CH3COO][64]. The efficiency in disrupting the crystalline structureof eucalyptus and sugarcane bagasse was observed to besuperior for [emim][CH3COO] than for [bmpy][Cl] [65].Cholinium amino acids ionic liquids were recently ap-
plied on rice straw pre-treatment and led to a substantialincrease of the cellulose CrI instead of the normal de-crease [66]. These contrasting effects on CrI resulted indifferences in the ability of these ILs to dissolve cellulosewhere the tested ILs are inefficient. Thus, an increase inthe CrI cellulose of the regenerated materials is onlycaused by a considerable removal of amorphous compo-nents, such as lignin, with no significant dissolution ofcellulose [66]. The regeneration method also influencesthe cellulose crystallinity where a vigorous stirring wasrequired. It may partially inhibit the induction of thecrystalline ordering within the regenerated sample andmay form an amorphous material [24].Furthermore, by treating switchgrass with [emim]
[CH3COO] at 160°C, Arora et al. investigated the sur-face morphology of switchgrass by measuring the BET(Brunauer-Emmett-Teller) surface area, and the porosityusing the BJH (Barret-Joyner-Halenda) method [67].Comparing to the untreated switchgrass, the BET surfacearea of the pre-treated switchgrass increased by a factorof 30. At the same time, the pore volume also increased30-fold with an average measured pore size of 10–15 nm
for [emim][CH3COO] of the pre-treated switchgrass[67]. Dibble et al. showed that a smaller particle size ofthe regenerated material was obtained by treating cornstover with the same IL [68]. Morphological changeswere also observed for pre-treated wood flour with[emim][CH3COO] and [bmim][CH3COO] [58]. This is agreat advantage of biomass pre-treatment with ILs, sincecellulose surface area decreases and thus, it providesgood accessibility to cellulase and enhances the catalyticefficiency of such enzyme [43,69].The regeneration process was explained by Zavrel
et al. describing that ions of the IL are extracted into theaqueous phase through hydrogen bonding, dipolar andCoulombic forces [26,70]. Basically, water moleculesform hydrodynamic shells around ions of the IL dis-rupting the direct interactions of IL ions with cellulose.Thus, intra- and inter-molecular hydrogen bonds are re-built and cellulose precipitates [26]. However, not onlycellulose can be dissolved but also other compounds oflignocellulose interact with the IL ions, such as hemicel-lulose and lignin. Singh et al. stated that the addition ofwater to switchgrass pre-treated in [emim][CH3COO]results in the precipitation of carbohydrates for down-stream processing and lignin appears to be partiallyextracted in the IL/water mixture [31]. The degree ofdelignification and interaction of lignin with the precipi-tating solvent, IL, or cellulose dictate the purity of theregenerated cellulose for further saccharification andsugar yield [31]. Leskinen et al. evaluated the precipita-tion and composition of the regenerated samples fromwood sawdust treatment with [amim][Cl] [35]. In theobtained regenerated material the lignin content in-creased due to the absence of delignification and simul-taneous dissolution of some polysaccharide components.However, Nguyen et al. presented that lignin is firstly re-moved in the pre-treatment of rice straw with [emim][CH3COO] [51]. This may indicate differences in themode of dissolution between [amim][Cl] and [emim][CH3COO] or it may also result from differences bet-ween biomass used in both studies. A more accuratecorrelation between lignin removal and the reduction ofcellulose crystallinity was approached in a work of Tanet al. [71] Lignin content in the regenerated material wasfound to be proportional to the CrI. Thus, when morelignin is retained in the regenerated material, higher CrIis observed. The possible explanation comes from themovement restriction of cellulose in the presence of lig-nin that causes the dissolved cellulose to rearrange intothe same conformation upon recrystallisation. Thus, thecrystallinity of cellulose remains relatively high if pre-treatment was not efficient in lignin removal [71]. For lig-nocellulosic materials that contain relatively large amountsof lignin, higher temperature is required to change thestructural crystallinity of cellulose I to cellulose II [59].
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In addition to the ability of an IL to reduce CrI andlignin content, other factors such as the promotion ofinter-crystalline swelling and reduction in fibre size mayplay an important role in determining the efficiency ofan IL as a pre-treatment solvent for lignocellulosic bio-mass [58]. Although the disruption of the cellulose crys-tallinity is crucial to provide the biomass suitable forenzymatic digestion, lignin removal is even more criticalfor improving enzymatic hydrolysis of cellulose [72].As mentioned above, the regenerated material is
mainly composed by cellulose, since this macromoleculeis not soluble in antisolvent. However, a 100% (w/w) re-generation of cellulose seems not to be possible. Duringthe pre-treatment process cellulose might undergo deg-radation and form smaller components soluble in the IL[71]. Moreover, the regeneration process is dependentupon molecular weight, where the dissolved higher mo-lecular weight and partially dissolved wood componentsin ILs are the first to precipitate [35]. The smallest mo-lecular weights could not precipitate after antisolventaddition being dissolved in the liquid stream.The choice of the antisolvent is also an important fac-
tor, which affects the yield of the regenerated wood.Li et al. studied the yield of regenerated wood with waterand methanol as antisolvents [56]. They determined thatthe regeneration yield was found to be higher in waterthan in methanol due to the easiness and strength of thehydrogen bond formation between antisolvent and bio-mass fraction [56]. Indeed, polar protic solvents such aswater, short-chain alcohols (methanol and ethanol) andeven acetic acid are able to precipitate efficiently thedissolved material [35]. In the case of using the relativelypolar chloroform as precipitating solvent, the efficiency ofregeneration was found to be good. However, consideringlimitation such as obstacles in treating waste streamscontaining chlorinated solvents and environmental issues,hazardous chemicals should be avoided. Acetonitrile canalso be used to precipitate biomass offering two advan-tages. The first benefit comes from its volatility and aneasy removal from the IL. The second advantage corre-sponds to the brighter fractions obtained by precipitationthat would allow for easier bleaching if required [35]. Aspreviously mentioned the reconstituted material presentsa gelatinous phase, thus it is hard to separate it from theviscous liquid after regeneration. Dibble et al. developedan antisolvent mixture with specific proportions of analcohol (ethanol) and ketone (acetone) that providedthe regeneration of solid precipitates with porous ap-pearance and easily dispersed in the liquid with no gel for-mation [68]. A rapid separation was attained and a lowervolume of antisolvent mixture was used in comparisonwith other antisolvent used. Mixtures of precipitating sol-vents can also be used in order to improve lignin extrac-tion yields with simultaneous high regeneration yields of
the carbohydrate fraction. Muhammad et al. suggested amixture of acetone and water in 7:3 ratio that reached al-most 100 wt% of lignin (indulin AT from Sigma Aldrich)solubility [42]. Wang et al. observed that addition ofDMSO to water as a precipitating solvent mixture in-creased the cellulose content in the regenerated materialin comparison with acetone/water mixture. Thus, DMSOwas reported to have greater lignin extraction capacitythan acetone [34].
The process condition effectsSpecific interactions between ILs and biomass are knownto be dependent on the IL, as well as on several conditionsinherent to pre-treatment, such as temperature, time, typeand particle size of biomass, solid/liquid ratio and watercontent. Major effects of these parameters in the biomasspre-treatment are described in the following sections,which contribute for a better understanding of pre-treatment with ILs.
Particle sizeThe particle size of biomass sample is one of crucial is-sues in lignocellulosic pre-treatment because such a par-ameter directly impacts on the contact and diffusion ofchemicals into the lignocellulosic materials [73,74].Kilpelainen et al. concluded that dissolution of wood
chips (size in excess of 5 mm × 5 mm× 1 mm) at hightemperatures in [bmim][Cl] and [amim][Cl] is complete[24]. Thus, smaller particles should be more suitable tobe used in the IL pre-treatment process [24]. The in-creased surface area of the smaller particles and the in-crease of mechanical pulping to breakdown the internalstructure are factors that improve the efficiency of bio-mass pre-treatment in ILs [25].Different responses are observed when different par-
ticle sizes of biomass are used. Sun et al. determined thesolubilities of different sized wood particles in [emim][CH3COO] and [emim][Cl] ILs [25]. The good dissol-ution efficiency of [emim][CH3COO] was preserved des-pite of the increase in the particle size of wood fromsmaller than 0.125 mm (99% solubility) to 0.5-1 mm(93% solubility). The different wood size solubilities weremore pronounced for [emim][Cl]. At wood size smallerthan 0.125 mm the dissolution reached 52.6% and withparticles size of 0.25-0.5 mm only 26.0% of dissolutionwas achieved. This suggests that [emim][CH3COO] ismore efficient and it would be much more economicalto use since the energy needed to reduce the biomassparticle size could be minimised [25]. Furthermore,Nguyen et al. investigated the influence of particle sizeusing the same IL in the pre-treatment of rice straw[51]. The study was conducted with three size ranges,smaller than 2 mm, 2–5 mm and larger than 10 mm de-termining the cellulose recovery and glucose conversion
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yields after pre-treatment. The treatment with the finestparticle size appears to have a considerable loss of cellu-lose content. Nevertheless, these samples showed thehighest glucose conversion, but it was ineffective due tolow cellulose content in the sample. The glucose con-version of the 2–5 mm particle-sized samples was sig-nificantly higher than that of > 10 mm particle-sizedsamples and similar to the value of lower than 2 mmparticle-sized samples [51].In a recent study, Bahcegul et al. proved that the ideal
particle size of a certain biomass for pre-treatmentmight change depending on the type of the IL used [64].Once again [emim][CH3COO] and [emim][Cl] wereused to compare the efficiency in the pre-treatment ofcotton stalks in four different particle sizes (< 0.15 mm,0.15-0.5 mm, 0.5-1.0 mm and 1.0-2.0 mm). They ob-served a decrease in the glucose yield for cotton stalkswith the smallest particle size (< 0.15 mm) in compari-son with larger particle sizes after pre-treatment with[emim][CH3COO]. This could indicate that particle sizeof biomass in pre-treatment with [emim][CH3COO]might influence the amount of cellulose recovery. Onthe other hand, [emim][Cl] works more efficiently withsmaller biomass particle sizes in respect to glucose yield.This opposite trend is explained by the different dissol-ution rates of the tested ILs. The [emim][CH3COO]showed a faster dissolution rate for the smaller particlesize allowing more extensive depolymerisation to lowermolecular weight water soluble compounds. Therefore, alower glucose yield was observed since part of cellulosewas retained as soluble compounds in the liquid phaseafter biomass regeneration. After enzymatic hydrolysis, itwas confirmed that different sizes of biomass could beideal for different ILs [64]. Bahcegul et al. also evaluatedthe lignin extraction after pre-treating the referred sizeparticles of cotton stalks [64]. For all tested particle sizesin the pre-treatment with [emim][CH3COO], less lignincontent was determined for each pre-treated sample.However, the highest lignin contents were observedfor larger biomass particle sizes (0.5-1.0 mm and 1.0-2.0 mm). Therefore, it is clear that pre-treatment with[emim][CH3COO] is more efficient using smaller par-ticle sizes. The same observation was not found in pre-treatment with [emim][Cl], since cotton stalk sampleswith different particle sizes had similar lignin contentafter pre-treatment [64].The crystalline structure presented by cellulose after
pre-treatment is also correlated with the biomass par-ticle size used. As referred above, transformation fromcellulose I to cellulose II could be observed using[emim][CH3COO]. In the pre-treatment of cotton stalkwith this IL, Bahcegul et al. demonstrated by XRD ana-lysis that this phenomenon could only be observed forthe smallest particle size studied (< 0.15 mm) [64].
Therefore, it can be inferred that smaller particle sizesare more appropriate to obtain pre-treated samples withhigher enzymatic digestibility.The pre-treatment efficiency is improved using small
particle sizes of biomass. However, size reduction of bio-mass for a pre-treatment process is questionable toachieve sustainability due to inherent high costs. Not-withstanding, ILs such as [emim][CH3COO] seem topreserve the pre-treatment efficiency for larger particlesindicating the possibility to reduce the energy demandon the grinding stage of biomass. Viell et al. studied thedisintegration and dissolution kinetics of differentparticle sizes of beech and spruce wood in [emim][CH3COO] [29]. They concluded that the dissolution issize-independent. They discovered that [emim][CH3COO]provided a quick swelling and disintegration of wood tis-sue. Consequently, this phenomenon creates a large spe-cific surface area which is favourable for the processing oflarger wood chips. Beech wood was dissolved more effect-ively and even larger wood chips of 10 mm length practic-ally exhibited the same dissolution kinetics as particleswith lower length [29].
Temperature effectGenerally, the temperature increase accelerates swellingand dissolution rates of lignocellulose in ILs [25,26,34,40].This phenomenon is possible due to destabilisation effectof temperature on the hydrogen bonds in the three-dimensional cellulose structure [26]. By decreasing tem-perature, longer times are required for an efficient swellingand dissolution of biomass [75]. Wang et al. describedsome physical aspects of wood chips dissolution in [amim][Cl] using a hot stage polarizing microscope [34]. Below50°C the wood samples had the same fibrous structure asits original state. The beginning of the swelling was no-ticed at 70°C and the dissolution occurred at 100°C. Thedissolution rate increased at 120°C and most of the fibrousstructure disappeared after only 10 minutes at thistemperature resulting in a slightly dark mixture. After30 minutes at 120°C no further changes were detected,and within a period of 4 hours, a fully black field observedindicated only a partial dissolution of the wood chips in[amim][Cl] [34]. Due to the importance of the reductionof the moisture in the mixture to enhance the pre-treatment process, temperatures above 100°C are generallyused. Muhammad et al. dissolved bamboo biomass in the[emim][Gly] IL containing 4.56 wt% of water, during6 hours at 120°C [42]. It was observed that the water con-tent decreased to 0.75 wt% after 350 minutes of heatingfacilitated the dissolution process of the bamboo biomass.The reduction of IL viscosity by increasing the tem-perature is also a factor favouring the swelling of lignocel-lulose particles and the mixing during the pre-treatmentprocess [43,71]. Indeed, the physico-chemical properties
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of the IL, such as viscosity and melting point, aretemperature dependent and govern conditions of the pre-treatment process in ILs. The [amim][Cl] as low viscosityIL can be used at lower temperatures. On the other hand,ILs with aromatic side chains such as [benzylmim][Cl] re-quire somewhat higher temperatures due to a high melt-ing point and higher viscosity [24,26].After dissolution the biomass regeneration is also af-
fected by temperature. In fact, some studies showedhigher regeneration yields at higher temperatures[43,76]. This is opposite to the statement that highertemperatures led to the compound degradation and con-sequently minor regeneration yields were obtained[34,41,71]. This controversy is explained by the differentmorphologies and compositions of the biomass used,which in turn present different responses to the tem-perature increase. A treatment of pine, using [amim][Cl]between 90°C and 120°C, showed the highest regener-ation rate at 120°C, but at the same temperature no re-generation product was obtained after catalpa treatmentindicating the occurrence of biomass degradation [34].Furthermore, pine was treated at 140°C and most of thedissolved biomass underwent degradation, similar to ca-talpa at 120°C [34]. However, without using much highertemperatures that cause biomass degradation, thecomplete dissolution in ILs might not be possible [35].Thus, the temperature applied in pre-treatment is de-pendent on the biomass and should be adjusted to anoptimal condition of pre-treatment.A study with sugarcane bagasse treated with [bmim]
[Cl] at different temperatures (110-160°C) showed thatbelow 150°C the crystallinity of cellulose-rich regene-rated product is slightly lower than that of the originalmaterial [76]. In all tested experiments no complete dis-solution was observed. However, temperatures above150°C demonstrated to reduce significantly the crystal-linity and to increase the saccharification rate andextent. Therefore, complete dissolution might not be re-quired to maximize saccharification efficiency at temper-atures above 150°C [76].The increase in temperature from 60°C to 120°C for 2-
hour pre-treatments of sugarcane bagasse with [emim][CH3COO] resulted in a water-soluble product thatrevealed a 10-fold magnitude extraction yield from theoriginal biomass and a 2-fold glucose yield in the enzym-atic hydrolysis of regenerated bagasse [69]. However, thexylose yield decreased as temperature increased from100 to 120°C indicating hemicellulose degradation inlow-molecular weight compounds extracted to the liquidphase [69]. Arora et al. used the temperature from 110to 160°C for a 3-hour pre-treatment of switchgrass usingthe same IL [67]. The obtained sugar yields increasedwith temperature allowing to achieve the best saccharideyield at 160°C, 12 times higher than that observed at
110°C. Hemicellulose was the most significant com-pound released from the pre-treatment process [67]. Itseems that higher temperatures expedite an autocatalyticprocess of hemicellulose which is accelerated for tem-peratures above 100°C [71], although this process is verydependent on the biomass and the IL used. On the otherhand, Labbé et al. described that at high temperatures,[emim][CH3COO] is capable to cleave the acetyl groupscovalently attached mostly to the hemicellulose com-ponent of yellow poplar [77]. Therefore, at highertemperature ILs could effectively disrupt the carbohy-drate–lignin linkages releasing hemicellulose.High temperatures were also beneficial for lignin ex-
traction in ILs. Tan et al. observed an improved delignifi-cation of sugarcane bagasse using alkylbenzenesulphonateIL ([emim][ABS]) with increasing temperature from 170°Cto 190°C [78]. The same trend was also confirmed inwood flour and triticale straw using [emim][CH3COO][28,79]. It is assumed that the most efficient delignificationtemperature is strongly related to the average glass transi-tion temperature of 165°C for a given lignin polymer [80].However, that temperature is dictated by the chemicalcomposition of lignin (monolignol composition and con-centrations) and varies significantly between the differenttypes of lignocellulosic biomass [67]. In the pre-treatmentprocess at higher temperatures, lignin is susceptible toself-condensation reactions. The condensation of lignin isaccompanied by an increase in molecular weight, whichresults in precipitation during the process and subse-quently lignin is recovered in the regenerated material.Moreover, inhibition of lignin dissolution after condensa-tion is another issue that contribute to the higher percent-age of lignin recovered in the regenerated material [81,82].Nevertheless, if higher temperatures were complementedwith long dissolution times, the condensed lignin couldstart to decompose and subsequently dissolve [83].Indeed, higher temperatures aid in improving the pre-
treatment efficiency, but at the same time results inmore degradation of carbohydrates and ILs themselves,as well as in modification of lignin properties. Moreover,the higher energy demand when using elevated tem-peratures is a drawback for sustainable and economicprocesses. Therefore, the choice of temperature for thepre-treatment process must be considered and optimiseddepending upon the biomass and IL used.
Reaction time effectThe pre-treatment duration time is related to the appliedtemperature in order to accomplish an efficient pre-treatment. It can be assumed that good results areexpected with a simultaneous short duration of time andhigh temperatures or using prolonged treatments at rela-tively low temperatures. Yoon et al. pre-treated the sug-arcane bagasse with [emim][CH3COO] and proposed a
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model based on Response Surface Methodology (RSM)to predict the reducing sugar yield by changingtemperature, time and biomass loading [43]. An im-provement in reducing sugar yield at longer pre-treatment at lower temperature (120°C) was noticed.However, prolonged pre-treatment could lead to de-crease in reducing sugar yield under higher temperatures(more than 135°C), explained by a depolymerisationprocess of the biomass components [43]. Sun et al. ob-served that complete dissolution of southern yellow pineand red oak in [emim][CH3COO] at 110°C was achievedafter 46 and 25 hours, respectively [25]. However, theimproved dissolution capacity provided by a longer pre-treatment time is inevitably accompanied by some deg-radation of either the dissolved biopolymers or the IL[25,67]. After ultrasonic pre-treatment of kenaf powderwith [emim][CH3COO] at 110°C, the hemicellulose con-tent decreases in the regenerated biomass to 6 wt% after120 minutes [50]. The reduced biomass recovery was at-tributed to partial hydrolysis of hemicellulose. Therefore,not only compound degradation occurs, but also the hy-drolysis of hemicellulose to reducing sugars that arethen lost in the washing stream [50]. Nguyen et al. dem-onstrated that an increase in the solubility time of ricestraw in [emim][CH3COO] caused a higher celluloserecovery, at least after 24 hours [51]. However, after48 hours of pre-treatment, the regenerated particleswere difficult to collect due to long time applied to pro-foundly break down cellulose chains from rice straw intosoluble oligosaccharides. There was no regeneration inamorphous cellulose form, thus decreased cellulose re-covery was observed [51]. In fact, the mean molecularweight of dissolved compounds was found to decreaseas the treatment time was extended [84], thus the com-pound regeneration was more difficult. Pre-treatment ofsugarcane bagasse with [emim][CH3COO] tends to forma coagulate at higher temperatures for a prolonged dur-ation time. This leads to a difficult regeneration of thedissolved sugarcane bagasse from the coagulate [43]. Sig-nificant amount of residual [emim][CH3COO] could beimprisoned in the coagulate, which might cause cellu-lase inactivation and consequently lead to a decrease inreducing sugar yield [85]. Furthermore, when higher tem-peratures are applied in biomass pre-treatment for ele-vated periods of processing, cellulose degradation is morepronounced [71,86]. Thus, shorter retention times arerecommended when high temperatures used, althoughthis must be balanced with the IL stability and potentialside reactions [40]. Tan et al. demonstrated that, fora shorter pre-treatment time, the glucose recoveryyield increased almost linearly from 26.2% to 96.7% asthe temperature increased from 60 to 100°C by pre-treating oil palm frond with [bmim][Cl] [86]. Neverthe-less, [emim][CH3COO] demonstrates more efficiency in
disrupting the lignocellulosic structure in a shorter periodof time, since it has a faster diffusion rate resulting inhigher glucose recovery yields [64]. The crystallinity de-gree of cellulose-rich material after regeneration was alsofound to decrease to an amorphous state as pre-treatmenttime increased [69].Long pre-treatment times were reported to favour lig-
nin extraction [66,71,78]. The diffusion of IL into thebiomass is improved and aids the dissolution and extrac-tion of lignin from biomass [71]. Lee et al. presented apre-treatment of wood flour with [emim][CH3COO]with varying time from 0.5 to 70 hours at 90°C [28].Around 40% of the lignin was extracted from wood flourwithin 5 hours and more than 85% was extracted after70 hours providing a cellulose-rich regenerated materialwith only 3.2% of lignin content for the last pre-treatment [28]. The same IL and temperature were ap-plied in pre-treatment of triticale straw resulting in21.1% of lignin extraction within 5 hours. An increase oflignin extraction yield up to 30.3% was noticed after24 hours of treatment [79]. The results from both stud-ies allow indicate that the lignin extraction is improvedas the pre-treatment time increases. However, the timerequired could be different regarding the biomass used.Viell et al. observed the effect of longer dissolution timeas more pronounced for beech wood than for sprucewood after pre-treatment with [emim][CH3COO] [29].This corresponded not only to a better dissolution rateof beech wood, but also to a higher lignin recovery [29].
Biomass type and concentrationThe degree of biomass recalcitrance varies as a functionof the biomass itself (i.e., grass, softwood, and hard-wood), and is influenced by inherent variations in termsof age, harvest method, extent of drying and storageconditions [62]. Furthermore, the lignocellulosic biomasscomprises different chemical and physical characteris-tics, such as composition of cellulose, hemicellulose andlignin, accessible surface area, crystallinity, degree ofpolymerisation, and others [41]. All of these featuresaffect the pre-treatment efficiency, thus special attentionis recommended to the type and concentration of bio-mass in the pre-treatment with ILs.Generally, softwood demonstrates lower density than
hardwood leading to much facile solubility in ILs [24].Pre-treatment of pine, poplar, Chinese parasol and ca-talpa wood chips with [amim][Cl] showed that pine washighly effective for pre-treatment providing high cellu-lose extraction yields, good dissolution and regenerationof pre-treated biomass. Furthermore, Li et al. comparedthe pre-treatment of three different biomass (southern pine,Norway spruce thermochemical powder and Eucalyptusgrandis) with [amim][Cl]. They concluded that underthe same conditions the efficiency decreased in the
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following sequence: southern pine > Norway sprucethermochemical powder > Eucalyptus grandis [56]. Thehigher density of Eucalyptus grandis leads to a more diffi-cult access of ILs reducing the pre-treatment efficiency[56]. Therefore, biomass species of lower density aredesirable. Thus for higher density biomass, more energyinput is required by increasing the temperature andtime of the process. However, Sun et al. found that redoak (hardwood) had a slightly higher dissolution in[emim][CH3COO] than southern yellow pine (softwood)[25]. This result can be associated with higher lignin con-tent in softwoods, which influences the dissolution, as wellas variations in the lignin-carbohydrate interactions, lignindistribution or lignin structure in hardwoods and soft-woods [25]. The different results are affected by the use of[emim][CH3COO] instead of [amim][Cl] highlightingthat the different properties of each IL can lead to differ-ent pre-treatment approaches regarding the nature of thebiomass used.The solid/liquid ratio (concentration of biomass) has
an influence on the dissolution capacity of ILs, regener-ation efficiency, enzymatic accessibility to biomass, glu-cose recovery and even on the conditions applied onpre-treatment process. Sun et al. studied a 4 to 10%(w/w) of wood load in [emim][CH3COO] at 110°C for16 hours and thus found an optimum concentration of5% (w/w) [25]. It was demonstrated that as the initialwood load increases, a lower percentage of the addedwood is dissolved [25]. In fact, low concentrations ofbiomass favour the dispersion of molecules in solutionand lead to high dissolution and regeneration rates.Wang et al. reported a decreasing trend in the dissol-ution amount from 35 to 26% when 1 and 5 wt% of ini-tial wood load in [amim][Cl] were tested, albeit a similardecrease was also observed for the same experiments[34]. The enzymatic accessibility of recovered cellulosecould be also affected by high solid/liquid ratio. Further-more, a higher solid loading can result in problems withagitation of the mixture and cause heat and mass trans-fer limitations in the mixture as well as a significantlydecrease in the IL accessibility to the biomass [71,86].These results show that pre-treatment in ILs could havelimitations with respect to the wood load capacity neces-sary to reach an efficient process. However, Tan et al.demonstrated that a high solid loading could improvethe pre-treatment efficiency [71]. This can be explainedby more frequent contact and collision between the bio-mass particles due to mixing at higher concentrations inviscous ILs that promotes cellulose dissolution. Yoonet al. optimise the conditions for the pre-treatment ofsugarcane bagasse in [emim][CH3COO] [43]. For this,the proposed optimal conditions were 145°C for 15 mi-nutes with a biomass loading of 14 wt%. They demon-strated a glucose recovery dependence on the overall
heat transfer in the mixture enhanced by frequent colli-sion of biomass particles [86]. Under higher biomassconcentrations, the chemical equilibrium of the reactionmight favour the cellulose dissolution at a higher tem-perature improving the glucose recovery [43]. Wu et al.accomplished relatively good efficiency in the pre-treatment of corn stover, maple wood flour, poplar andswitchgrass with [emim][CH3COO] using solid/liquidratio of 33% (w/w) [41]. They described that, at thisspecific biomass loading, the molar ratio of [emim][CH3COO] to glucose in cellulose (6:1) is comparable tothe possible number of hydrogen bonds (acceptor anddonor) for each glucose unit in cellulose. With this stoi-chiometry there are enough [CH3COO] anions to inter-act with the hydroxyl groups of each glucose unit and todisrupt effectively the crystalline structure of celluloseduring the pre-treatment process. If the biomass concen-tration was higher, the crystalline disrupting of cellulosewould be incomplete and thus the efficiency of the pre-treatment could decrease [41]. When the concentrationof biomass compromises the pre-treatment efficiencydue to a high viscosity of the moiety, ultrasound irradi-ation seems advisable to be used. Such a solution isintended to improve the diffusion of ILs into the bio-mass to attain an efficient disruption [50].Notwithstanding, the increase of biomass concentra-
tion in ILs is a challenging task for process intensifica-tion (to reduce the energy demand, as well as the size ofthe equipment) therefore the pre-treatment process opti-misation is needed.
Water presence influenceIn the dissolution process, careful attention to watercontent should be considered originated either from theIL or from the natural biomass moisture. The water con-tent in ILs (above 1 wt%) was found to reduce the solu-bility of cellulose [19]. Less than 1 wt% water content isreliable for efficient cellulose dissolutions by ILs. Waterconcentration higher than that provides enough watermolecules to compete with IL anions to form hydrogenbonds with cellulose thus promoting only partial dissol-ution of lignocellulosic biomass [19,24]. In the case ofusing chloride-based ILs, the presence of water shouldbe more restrictive to not coordinate with chloride ions[48]. However, a certain amount of water seems to benecessary to promote lignin dissolution in [bmim][Cl][40]. On the other hand, the presence of water in theacetate-based ILs reduces their hydrogen bond acceptorcapability thus compromising the dissolution efficiency[58]. Therefore, ILs should be dried prior performingbiomass pre-treatment. If the biomass used is air-dried,a greater amount of water content would be introducedinto the dissolution process thus decreasing the solventcapacity of ILs and subsequently the pre-treatment
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efficiency [40,87]. For dried biomass a rigorous dryingprocess should be applied, which is demanding in costand energy.
The pre-treatment of biomassThe subject of the biomass processing using ILs is veryrecent. The pre-treatment with ILs enables one to: (i)alter the physicochemical properties of the biomassmacromolecular components; (ii) extract a specific ma-cromolecular component that is provided by the pro-perty of ILs; and (iii) perform different fractionationapproaches after biomass dissolution in ILs. Main condi-tions and results of biomass pre-treatments in ILs arecompiled in Tables 1, 2 and Additional file 1: Table S1.The pre-treatment is dependent on the IL, lignocellu-losic biomass (type, moisture, size and load), tempera-ture, time of pre-treatment and precipitating solvent used.In this section the advances of the IL pre-treatment tech-nology are intended to be described in detail including apositive impact on the maximal exploitation of lignocellu-losic biomass concerning the biorefinery concept. The
Figure 3 Schematic representation of the pre-treatment of biomass uhemicellulose and lignin. Saccharification and fermentation processes canand hemicellulose. Valuable products can also be generated by processingbiorefinery process.
Figure 3 depicts the pre-treatment of biomass using ILs,the further fractionation and downstream processes.Until now, [emim][CH3COO] seems to be the most
suitable IL for the pre-treatment of lignocellulosic bio-mass, since it possesses good solvent power for thesematerials and hence it is also referred to in the moststudies of this research field. Several studies were per-formed focused on the evaluation of the pre-treatmentbehaviour of different hardwood and softwood specieswith [emim][CH3COO] [25,28,29,32,58,88]. Sun et al.performed a 5% (w/w) dissolution of southern yellowpine in [emim][CH3COO] (110°C, 16 hours) and ap-proximately 59% of the original carbohydrates was re-covered in the regenerated material after adding anacetone/water mixture as the antisolvent [25]. Addition-ally, around 31% of the original lignin was obtained infree form and 38% linked to carbohydrates as part of thereconstituted wood, leading to a lignin reduction of26.1%. The same treatment was also performed forred oak with a 34.9% reduction of lignin was reached.Hence, the regenerated materials contained less lignin
sing ILs and further regeneration and fractionation into cellulose,be integrated to produce biofuels and bioproducts from cellulosecellulose, hemicellulose and lignin fractions within the
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than the original wood, with a fairly good delignificationat relatively low temperature reported. However, sig-nificant losses of carbohydrates and lignin during thefractionation process were observed. In the case of pinepre-treatment, total losses of 41% and 31% of the originalcarbohydrates and lignin were calculated, respectively[25]. Lee et al. extracted approximately 63% of the initiallignin from the wood flour, at 130°C after 90 minutes oftreatment using the same IL [28]. However, they alsoextracted 16% of the cellulose and 26% of the xylan fromthe initial wood flour content. Therefore, the processshowed significant delignification with small losses of car-bohydrates after the pre-treatment [28]. In a differentstudy, by the temperature decrease to 90°C and the timeof pre-treatment increase to 24 hours for the samebiomass loading, nearly 50% of the lignin was ex-tracted from the initial wood flour sample [58]. Such a re-sult confirms a great effect of the applied conditions(temperature and time) in the pre-treatment of lignocellu-losic biomass. Torr et al. evaluated the pre-treatment ofcompression pine wood (Pinus radiata) at 120°C and155°C for 3 hours [30]. When water was used as theantisolvent, no lignin extraction was observed. Thesefindings were the opposite to those found earlier by Sunet al. [25]. The authors justified different results as a con-sequence of the antisolvent used. The reported pre-treatment removed mainly hemicelluloses for both testedtemperatures leading to the recovery of a pre-treated ma-terial with high cellulose and lignin contents. With an ex-tensive study of the pre-treated biomass using HSQCNMR technique, Torr et al. verified the existence ofdeacetylation of hemicellulose, possible depolymerisationof polysaccharides, loss of lignin ether linkages and for-mation of lignin condensation products [30].Pine, poplar, Chinese parasol and catalpa wood pow-
ders were pre-treated using a solvent mixture of [amim][Cl]/DMSO at the optimised conditions of 110°C for2 hours and a 3 wt% wood concentration under micro-wave irradiation [34]. In each sample, wood chip/IL sus-pension was prepared by the addition of 16 wt% DMSOto reduce the viscosity of the mixture and facilitate fil-tration in biomass processing. The addition of DMSOhad no obvious impact on the solubility of the woodchips in the IL. Better results were reported for pinepre-treatment, in which more regenerated material wasobtained (56%) and the cellulose content reached 85%,being nearly 20% higher than that in the regeneratedpoplar, Chinese parasol and catalpa chips [34]. This re-sult is explained by a higher solubility of pine wood in[amim][Cl] that provided the best performance in thepre-treatment. The original pine wood was composed of49% cellulose and the extraction rate of this pre-treatment could reach 62% of the total cellulose. Afterpre-treatment, a 14% loss of the original wood was
registered. Wang et al. described that biomass loss oc-curs from water-soluble low molecular matter that re-sults from degradation of wood, washing and filtrationsteps during the process [34]. Vo et al. also reported apre-treatment of poplar wood (5 wt%), but with [emim][CH3COO] at 130°C for 12 hours, achieving a decreaseof a 30% lignin content of pre-treated biomass relativeto the original content [32]. Other pre-treatments withwoods including poplar, oak, fir and acacia wereperformed with [emim][CH3COO], [emim][COOCF3],[emim][BF4] and [bmim][PF6] ILs at 90°C for 24 hourswith 2% (w/w) solid/liquid ratio [88]. The work of Voet al. demonstrates that [emim][CH3COO] had the bestperformance, since the total carbohydrates of the pre-treated biomass were completely hydrolysed by cellulase[88]. The compiled data of pre-treatments using woodbiomass, namely softwood and hardwood are presentedin Tables 1 and 2, correspondingly.Arora et al. showed that a 3-hour pre-treatment of
switchgrass at 160°C with [emim][CH3COO] resulted indelignification of 73.5% [67]. The regenerated solid iscomprised of 31% glucan (80% of the original glucanfrom switchgrass) with 8% glucan (20.5% of original glu-can concentration in switchgrass) in the liquid fraction.In contrast, the majority of the xylan (19%) ended up inthe liquid stream, which was 73% of the original xylanconcentration of switchgrass. This means that the de-polymerisation of xylan occurred and by HPAEC ana-lysis, it was concluded that depolymerisation resulted inthe formation of oligosaccharides instead of monosac-charides. Only 5% of lignin in the regenerated solid wasdetected [67]. Therefore, as stated previously, in thepresence of IL, higher temperature favours hemicellulosehydrolysis when using [emim][CH3COO] in the pre-treatment. Using the same biomass (from the same origin)and IL, as well as the same conditions of pre-treatment,Li et al. [62] showed different results. The pre-treatmentprocess partially removed lignin and hemicellulose fromthe original switchgrass and produced a regenerated ma-terial composed by 67.7% glucan, 13.6% lignin and 7.6%xylan content. The lignin extraction achieved 69.2% oftotal lignin removal including 12.0% of acid soluble lig-nin and 57.2% of Klason lignin [62]. Much better resultswere obtained by Arora et al. [67] They were explainedby a prior to and extended pre-extraction of switchgrassusing ethanol comparing to those performed by Li et al.[62] When switchgrass was pre-treated with [Me(OEt)3Et3N][CH3COO] at 110°C for 15 minutes, the content ofKlason lignin in the regenerated material was reducedto 24 wt% from the initial value of 34 wt% [89]. Al-though [Me(OEt)3Et3N][CH3COO] presented an im-proved solubility of alkali lignin as compared to [emim][CH3COO] [89], the bulky cation [Me(OEt)3Et3N] wasdemonstrated to have lower efficiency in the switchgrass
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pre-treatment in comparison with the smaller [emim]cation.The pre-treatment of crop residues were also per-
formed using different ILs. The [emim][CH3COO] wassuitable for a 48-hour pre-treatment of rice straw pro-viding 82% of cellulose recovery with subsequently 88%conversion to glucose after 130°C [51]. The same bio-mass was treated for 5 hours at 120°C with [bmim][CH3COO] and lignin content was reduced by 8.2%. Xy-lan content was also decreased from 21.7% for untreatedrice straw to 15.1% for the pre-treated biomass [90]. Ricestraw was pre-treated using the amino acid-based ILnamely, [Ch][Lys], [Ch][Gly], [Ch][Ala], [Ch][Ser], [Ch][Thr], [Ch][Met], [Ch][Pro] and [Ch][Phe] [66]. In allcases, the content of cellulose increased from 37.7% inthe original biomass to 48–62% after pre-treatment andthe content of acid-insoluble lignin decreased from16.9% of original lignin content to 10–12% in the re-generated material (depending on the IL used). A strongcorrelation between enzyme performance and delignifi-cation of rice straw was observed. A total of 94% and65% cellulose and xylan digestibility, respectively, wereobtained after pre-treatment with [Ch][Lys], [Ch][Gly],[Ch][Ala], and [Ch][Met] that were the most efficient inlignin extraction (>55%) [66]. Cotton stalk was subjectedto pre-treatments with several ionic liquids such as[HEA][HCOO], [amim][Cl], [bmim][Cl], [emim][Cl] and[emim][CH3COO]. The extracted lignin ranged between31% and 46%, where maximum removal yields wereobtained for pre-treatments with [emim][CH3COO] and[amim][Cl], determined to be 45% and 46%, respectively[63]. Kenaf pre-treatment was additionally performedwith five types of ILs, such as [amim][Cl], [bmim][Cl],[emim][Cl], [emim][DEP] and [emim][CH3COO] [50].The cellulose saccharification ratio was about 20% forkenaf powders pre-treated in [amim][Cl], [bmim][Cl],[emim][Cl] and [emim][DEP] by conventional heating at110°C for 2 hours. The glucose yield was further in-creased when ultrasonic radiation was used for the sameILs with a 60-95% glucose yield after 120 minutes atonly 25°C. The cellulose saccharification ratio of kenafpowder in [emim][CH3COO] was 86% after 15 minutesof ultrasonic pre-treatment at 25°C, compared to only47% in the case of the thermal pre-treatment in thesame IL. However, it seems that the ultrasonic pre-treatment leads to a higher loss material in the liquidphase than for the conventional oil heating pre-treatment [50]. Wu et al. reported a 33% (w/w) loadingof corn stover pre-treated with [emim][CH3COO] as anefficient pre-treatment, in terms of the biomass loading,since it removed 15% of the total lignin from the originalbiomass [41]. The best result was obtained by a pre-treatment with a biomass loading of 4.8% at 125°Cfor 1 hour giving a lignin extraction of 43.5%. The
regenerated material was determined to compose a63.4% carbohydrate and still remained 13.3% lignin [41].At 130°C wheat straw was pre-treated with [emim][DEP](4% w/w) resulting in a porous and amorphous re-generated material, with only 54.8% of reducing sugaryield attained possibly due to only a 30-minute pre-treatment process [91]. The dissolution of sugarcane ba-gasse in [bmim][Cl] at high temperatures, such as 140°Cand 150°C, was reported to be incomplete [76]. The un-dissolved material was first separated before regenera-tion of the dissolved material, the composition of whichwas practically the same as that of the original biomass.The results showed a composition of a regenerated ma-terial (without undissolved biomass) of 77% cellulose at140°C and 82% at 150°C. The lignin and hemicellulosecontent proportionally decreased as the temperature in-creased. At 150°C, the lower lignin and hemicellulosecontent of the regenerated material can be attributed toboth preferential dissolution of cellulose and to an in-creased degradation of hemicellulose and lignin duringthe pre-treatment process. The pre-treatment efficiencywas then evaluated by enzymatic hydrolysis and the bestresults were obtained for a pre-treatment at 150°C for90 minutes [76]. Sant’Ana da Silva et al. demonstratedthat only a 30-minute sugarcane pre-treatment at 120°Cwith [emim][CH3COO] gave a regenerated biomass verysusceptible for saccharification [69]. Yields above 95%and 76% of glucose and xylose were achieved, respect-ively. The [emim][CH3COO] IL was mixed with energycane bagasse at a 20:1 ratio [92]. It was observed that15.1% of the total mass was lost during pre-treatmentwith IL and 52.6% of the loss was attributed to lignin re-moval. Composition of analysis revealed that 32.1% ofthe initial lignin was removed in IL-treated energy canesamples. Specifically, 41.7% of the initial acid insolublelignin was removed from energy cane. The loss of glucanin IL-treated samples was less than 9% and cellulose di-gestibility of pre-treated sample at 24 hours was approxi-mately 25 and 22 times higher than untreated andwater-treated samples, respectively. The hemicellulosedigestibility followed the same trend as that observed forcellulose [92]. Tan et al. used oil palm frond that waspre-treated with [bmim][Cl] at different pre-treatmentconditions [86]. The pre-treatment variables studiedwere temperature, time and solid loading. Under thepre-treatment conditions of 4 wt% solid loading at 100°C,the lignin removal efficiency reached its maximum inless than a half hour. The lignin content decreased from33.63% in original biomass to 17.91%, i.e., close to 50%of the lignin was extracted. During the pre-treatmentprocess the cellulose loss was determined to be around4-9% [33]. Indeed, ILs are able to depolymerise polysac-charides in biomass to compounds with low molecularweight. ILs with greater depolymerisation power cause
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greater loss of reducing sugar during pre-treatment pro-viding higher mass loss of carbohydrates. The dissolvedpolysaccharides undergo different degrees of depolymer-isation and reducing sugar loss when pre-treated withdifferent types of ILs. Sugarcane bagasse was pre-treatedwith [bmim][Cl] and 6.7 mg · mL-1 of reducing sugarswas detected in IL. In the case of the pre-treatment with[emim][DEP], 2.7 mg · mL-1 of reducing sugars weredetected in IL, whereas [emim][CH3COO] was ineffect-ive in depolymerisation of polysaccharides during pre-treatment (insignificant amount of reducing sugar wasdetected) [33]. Tan et al. claimed that depolymerisationof cellulose occurs under acidic conditions in the ionicliquid media during pre-treatment [71]. The acidic con-ditions are created by degradation products, namelyacetic acid and carboxylic acid released from hemicellu-lose hydrolysis upon pre-treatment. Under acidic condi-tions IL had the ability to hydrolyse cellulose causingsignificant losses. The loss products are essentiallyformed by carbohydrate oligomers and monomers thatare soluble in the antisolvent and failed to be rege-nerated [71]. Van Spronsen et al. presumed that water isformed at temperatures higher than 100°C from dehy-dration [27]. The components of lignocellulosic biomasswere then exposed for hydrolysis. Hemicellulose couldbe further hydrolysed into acetic acid, which subse-quently may act as a catalyst to the hydrolysis process ofcellulose and the remaining hemicellulose [27]. Thelesser depolymerisation observed for [emim][CH3COO]could be explained by the buffering effect exhibitedby a very basic [CH3COO] anion. Its ability to com-bine with protons to form acetic acid may limit theacid-catalysed hydrolysis of hemicellulose polymers[53]. If the resultant products of degradation and hy-drolysis are low macromolecular weight compounds, theprobability of its recovery in the regeneration process islower. Additional file 1: Table S1 presents the pre-treatment results for grasses, agricultural and forest resi-dues, and industrial residues.The extraction and recovery of lignin are also in evi-
dence in some pre-treatment studies using ILs with themain goal envisaged for lignin valorisation. Tan et al.studied treatment at atmospheric pressure and hightemperature (170–190°C) with the IL containing [emim]and alkylbenzenesulphonate and xylenesulphonate anions[78]. They demonstrated that lignin can be extracted fromsugarcane bagasse with high yields. Lignin was recov-ered from the IL by alkali extraction and a yield oflignin above 93% from the original lignin was obtained.NaOH aqueous solution as the antisolvent was usedleading to the simultaneous extraction of lignin. Thecellulose-rich pulp obtained (with hemicellulose con-tent) corresponded to a 45–55% (w/w) from the originalsugarcane bagasse. However, the possible formation of
lignin carbonium ions during the heating dissolutionprocess was noted. The formed carbonium ions wereable to react with the xylenesulphonate anions, andthus lignin with xylenesulphonate adducts were pro-duced. Additionally, during delignification some amount ofhemicellulose was also removed [78]. In the pre-treatmentof triticale straw using 5% (w/w) of biomass treated at 90°Cfor 24 hours under N2, ILs, such as [emim][CH3COO],[bmim][Cl], N,N-dimethylethanolammonium formate([DMEA][HCOO]), N,N-dimethylethanolammonium acetate([DMEA][CH3COO]), N,N-dimethylethanolammoniumglycolate ([DMEA][OHCH2COO]) and N,N-dimethylethanolammonium succinate ([DMEA][(CH2COO)2] wereevaluated [79]. Lignin extraction was performed more effi-ciently by [emim][CH3COO] and [bmim][Cl]. The otherfour ILs, [DMEA][HCOO], [DMEA][CH3COO], [DMEA][OHCH2COO] and [DMEA][(CH2COO)2] were observedto have poor extraction performances. The extraction oflignin with [emim][CH3COO] was 30.3% of the originallignin from the triticale straw. This result was 2 and 10-fold higher than for [bmim][Cl] and [DMEA][HCOO] lig-nin extractions, respectively. The [bmim][Cl] IL is lessefficient than [emim][CH3COO] for delignification ofstraw. Additionally, [DMEA][HCOO], [DMEA][CH3COO],[DMEA][OHCH2COO] and [DMEA][(CH2COO)2] are notsuitable for this purpose [79]. The pre-treatment with[emim][CH3COO] also revealed a small change in the cel-lulose content in the regenerated biomass. Partial degrad-ation of xylan led to a loss of approximately 7% celluloseand 31% xylan after pre-treatment. Wheat straw and flaxshives were also pre-treated with [emim][CH3COO] underthe same conditions and it was identified that 29.6% and14.0% of lignin was extracted, respectively, from these bio-masses [79]. One of the most important conclusions com-ing from this study is the fact that the type of biomassaffects the extraction of lignin strongly, mostly due to thedifferences in composition and structural interactions oc-curring in various biomasses. Muhammad et al. presented[emim][Gly] as an effective solvent for delignification ofbamboo biomass due to a significant lignin reductionof 85.3% of total lignin content. The pre-treatment wasperformed at 120°C for 8 hours (5% w/w) [42]. Use ofacetone/water mixture (7/3 vol/vol) as the antisolvent wasfound to be advantageous due to the cellulose precipita-tion and a high lignin extraction yield obtained [42]. Otherstudy focused on the lignin isolation was made byShamsuri et al. who used the [mmim][MeSO4] IL to ex-tract lignin from rubber wood [61]. Depending on theconcentration of the IL, the time and temperature of thepre-treatment varied. The best concentration for thehighest lignin yield was found at 0.5 moles at 100°C after120 minutes with 0.224 g lignin soluble. At these condi-tions, it was possible to isolate only 13.03 wt% lignin ofthe rubber wood. This result indicates that [mmim]
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[MeSO4] is not efficient in lignin extraction from rubberwood. There is a large loss of rubber wood lignin in thewaste liquid (after lignin isolation), possibly due to its highwater solubility and/or its relatively small molecule di-mensions remained in the waste liquid. However, the iso-lated lignin was demonstrated to be nearly pure by FTIRstudy [61]. A relatively low extracted lignin content fromwood flour was obtained with the [bmim][MeSO4] treat-ment. On the other hand, [bmim][MeSO4] was capable ofdissolving a high concentration of commercial free lignin.In the case of [bmim][MeSO4], insufficient lignin extrac-tion from wood flour could be explained by a limited ac-cess of the IL to lignin in the wood flour causing arestricted extraction [58]. Considering pre-treatmentof rice straw, recent renewable amino acid-based ILs,such as [Ch][Gly], [Ch][Phe] and [Ch][Lys] were foundto favour the lignin extraction at 90°C [66]. [Ch][Gly]removed 59.5% of the total lignin, while [Ch][Phe]extracted only 41.5%. Surprisingly, [Ch][Lys] showed thehighest capacity to extract lignin from rice straw giving a60.4% yield, in spite of a moderate solubility of lignin in[Ch][Lys] [66].The next direction in studies on the biomass pro-
cessing is focused on the extraction of carbohydratefractions using ILs. Abe et al. attempted to use short dis-solution time (up to 2 hours) and low temperature (25°Cand 50°C) in the extraction of carbohydrates from branusing phosphonate or phosphinate-derived ILs [52]. Cel-lulose, hemicellulose, and residual starch were the majorcomponents dissolved by these ILs. The insoluble por-tion was expected to contain lignin and its complexes.The polar and low viscous [emim][PO(O)H2] IL waschosen among the investigated ILs for this study. A 42%yield of the extraction of carbohydrates present in branwas obtained at very mild conditions of 50°C within2 hours [52]. Another example of extraction of carbohy-drates was presented by Pezoa et al. who examined pre-treatment of eucalyptus and lenga residues using [emim][Cl] [60]. They achieved a carbohydrate extraction of 46-48% from the total carbohydrates present in wheatand corn residues, after a pre-treatment at 150°C for60 minutes. In the case of eucalyptus, the same con-ditions gave a 33% yield of total sugars. However, thelenga pre-treatment at a temperature of 150°C and anincubation time of 30 minutes yielded 40% of the ori-ginal carbohydrates. At 170°C a carbohydrate degrad-ation was also observed that is not beneficial if thegoal is to preserve carbohydrates for subsequent en-zymatic hydrolysis [60].When different conditions are applied to the pre-
treatment process, a multivariate analysis is advisable toobtain predicted conditions that ensure maximum ef-ficiency for a certain biomass pre-treatment with theparticular IL. Yoon et al. used the statistical RSM for the
operating conditions of pre-treatment of sugarcane ba-gasse with [emim][CH3COO] [43]. Temperature, timeand solid loading were explanatory variables tested andthe reducing sugar yield (enzymatic hydrolysis) was theresponse variable. As it was already stated for sugarcanebagasse pre-treatment in [emim][CH3COO] conditionswere optimised to be 145°C, 15 minutes and 14 wt% ofbiomass loading. At these conditions, a predicted redu-cing sugar yield was 71.0%, but the experimental redu-cing sugar yield reached 69.7%. However, consideringdelignification, this optimised pre-treatment was inef-fective. The crystallinity of cellulose was altered ratherthan the lignin removal during pre-treatment. Part ofthe lignin suffered from modification to a more acid sol-uble type after pre-treatment being precipitated togetherwith carbohydrates in the regenerated material [43]. An-other RSM study allowed to determine the optimisedconditions in the case of oil palm frond pre-treatmentwith [bmim][Cl] by evaluation of the glucose yield [86].The theoretical optimal conditions were determined tobe 80°C, 15 minutes retention time and a 10% (w/w)biomass loading. This optimised result gave a reductionof 13% and 38% in terms of temperature and time ofpre-treatment, respectively, as well as an increase of 19%biomass loading, in comparison with the best resultattained from the experimental work. The theoreticaloptimised conditions were tested and the predictedvalues from RSM study were validated obtaining a 100%glucose yield after enzymatic hydrolysis [86]. The RSMstatistical approach seems to be advisable to performoptimised pre-treatments of a certain biomass with aspecific IL.ILs apart from being used in the biomass pre-
treatment, can also be applied as a tool for the fraction-ation of lignocellulosic materials into the three maincomponents (Table 3). In a recent fractionation ap-proach cellulose, hemicellulose and lignin were obtainedseparately as solid fractions [93]. A sugarcane bagassepre-treatment was performed with 2% (w/w) of sugar-cane bagasse at 110°C for 4 hours. The complete dissol-ution of biomass in [bmim][Cl] achieved followed by theaddition of acetone/water (9:1, v/v) as antisolvent. Acet-one soluble lignin was extracted to the liquid streamwhereas cellulose, hemicellulose and alkaline lignin werefractionated from the solid fraction using a 3% NaOHsolution and ethanol. The biomass was fractionated to36.78% cellulose, 26.04% hemicellulose, and 10.51% lig-nin, giving 47.17% and 33.85% of the original polysac-charides and 54.62% of the original lignin. High purityfractions were obtained for cellulose (92.15% glucose)and even for the hemicellulose fractions (73.25% xylose,8.31% arabinose and 16.58% glucose). The glucosecontent in hemicellulose is derived from the cellulosehydrolysis during pre-treatment in IL and alkaline
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extraction [93]. A different fractionation approach wasperformed using [amim][Cl] in the pre-treatement ofbamboo biomass [94]. After a 12-hour pre-treatment at100°C, water was used to regenerate the dissolved bio-mass. The regenerated material was separated from theliquid phase containing water, IL and non-regeneratedbiomass. A subsequent two-step alkaline extraction ac-companied by an ethanol selective fractionation wasperformed. Cellulose-rich material, two fractions ofhemicellulose and two fractions of lignin were obtained.The cellulose-rich fraction demonstrated high cellulosecontent attaining 92.02 wt% purity. On the other hand,the lignin fractions presented traced amounts of carbo-hydrates (1.43 wt%-2.80 wt%) demonstrating high effi-ciency of the fractionation. Furthermore, the alkalineextraction led to the separation of hemicellulose frac-tions with different content in xylose and arabinose. Amajor content of arabinoxylan was obtained in onehemicellulosic fraction (80.52 wt% xylan and 7.04 wt%arabinan) while another fraction had an enriched con-tent of glucan derived from cellulose (52.97 wt% xylanand 42.75 wt% glucan). However, only 40.26% (w/w)of the original biomass was recovered resulting in 60%(w/w) material loss being the major limitation of thepresented process. Moreover, degradation of lignin andhemicellulose during the pre-treatment with [amim][Cl]promoted not only high material loss, but also increasein the polydispersity of the fractionated samples [94].Another literature report that provides informationabout fractionation was designed for pulverised andcoarse Norway spruce (Picea abies) and Eucalyptusgrandis wood dissolved in [amim][Cl] [35]. Leskinenet al. demonstrated fractionation based on the molecularweight of macromolecules [35]. In this study the select-ivity of precipitation was found to be not significantlydependent upon antisolvents. The selectivity of the frac-tionation was also dependent on the biomass used, aswell as on type of milling used before pre-treatment with[amim][Cl]. Cellulose was the first to precipitate as adelignified and bleachable macromolecule. The groupalso examined water extraction of the high molecularweight acetylated galactoglucomannan (hemicellulosiccompound). The major advantage of an effective millingand ball-milling before pre-treatment with ILs is the dis-integration of extensive lignin-carbohydrate complexthat allows the separation of lignin and polysaccharides[35]. A selective fractionation was also performed bypre-treating rice hulls with several ILs, such as [emim][CH3COO], [amim][Cl] and [hmim][Cl] [95]. After thedissolution process, water was added and the regenera-ted material was recovered. The filtrate was further usedand ethanol was added to give a second precipitatedmaterial. The pre-treatment with [emim][CH3COO] at110°C for 8 hours was found to completely remove
lignin from rice hulls and lignin was recovered in a sec-ond precipitation using ethanol as an antisolvent. Underthe same conditions with [hmim][Cl] used, approxi-mately 20% of the cellulose in the rice hull sample canbe precipitated out using water as an antisolvent, whilemore than 60% of the hemicellulose can be precipitatedwith ethanol [95]. The pre-treatment with [amim][Cl] atthe abovementioned conditions was demonstrated to re-move about 75% of hemicellulose content from ricehulls. Furthermore, in this fractionation approach, ashwas also removed and quantified. The pre-treatmentwith [hmim][Cl] removed 10-20% of the ash and there-fore provided the highest removal among the tested ILs[95]. Basing on the provided literature, it can be con-cluded that different fractionation approaches can be ap-plied using the specific properties of ILs to achieve aprecise goal in the biomass processing.The pre-treatment with ILs is not only more selective
for biomass fraction separation, but also can be consid-ered as more economic with respect to the energy con-sumed in the performance of the whole process. Yoonet al. evaluated the energy consumption of the pre-treatment with [emim][CH3COO] (145°C, 15 minutes,14% (w/v) of biomass loading) in comparison with alkali(115°C, 38 minutes, 9% (w/v) of biomass loading) andacid (150°C, 30 minutes, 15% (w/v) of biomass loading)pre-treatments [33]. The authors calculated a theoreticalmaximum yield of reducing sugars of 55.6 ± 0.2% forpre-treatment with [emim][CH3COO], and 57.2 ± 0.3%and 22.2 ± 0.4% for alkali and acid pre-treatments, re-spectively. Considering these theoretical values, the pre-treatment with [emim][CH3COO] consumes 1262.3 J. Inthe cases of the acid and alkali pre-treatments a total en-ergy was calculated to be 1847.9 J and 2611.8 J, cor-respondingly. It was also determined that 1.80 MJ isnecessary to pre-treat 1 Kg of sugarcane bagasse with[emim][CH3COO], in contrast to higher values of 4.11MJ and 3.48 MJ in the cases of alkali and acid pre-treatments, respectively. These results indicate that bio-mass pre-treatment with ILs can be more energeticallyefficient than other conventional pre-treatments, such asacid and alkali pre-treatments [33].
The biomass pre-treatment with aqueous-IL mixturesA novel approach in the pre-treatments with ILs washighlighted recently [53,55,58,96,97]. The principle aimis water addition to the dissolution process, on the con-trary to Swatloski et al. who reported that water com-promises the dissolution efficiency of biomass in the ILdissolution [13]. The use of aqueous-IL solutions wasfound to be advantageous as it led to: (i) a smalleramount of IL to be used; (ii) easier processing operationsdue to the reduced viscosity; and (iii) facilitated recyclingof the mixture instead of pure IL as requirement of the
C – cellulose; H – hemicellulose; L – lignin; O – others; a dried at 105°C for 24 h; b dried at 105°C oven for at least 24 h; c extraction yield; d dried for 16 h at 60°C; e glucan basis; f xylan, arabinan and galactan basis;g xylan, arabinan, mannan and galactan basis; h dried for 16 h at 55°C; i xylan, arabinan, galactan and rhamnan basis; j 87% total glucan; k 66% total xylan and arabinan; l 35% total lignin; m 14% total xylan; n 17% totallignin; o 5.9% total lignin.
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separation of IL and water could be omitted as reducingcosts of the pre-treatment [96,97].One of the first works in this subject was performed
by Doherty et al. who blended 5 and 10% (w/w) of waterwith [emim][CH3COO], [bmim][CH3COO] and [bmim][MeSO4] to pre-treat wood flour [58]. After a 12-hourpre-treatment at 90°C, a decrease in glucose and xyloseyields was observed with increased addition of water.The added water displayed a moderate inter-crystallineswelling effect and the wood fibres were still largely in-tact. The crystallinity and lignin content of the pre-treated wood flour were verified to be higher than thatof pre-treatments with pure ILs. In fact, the addition ofwater decreased the ability of the ILs to disrupt thecrystallinity of cellulose present in wood, affected bothintra- and inter-crystalline swelling, as well as fibre sizereduction. The lignin extraction varied from 21% to 24%(w/w) for all pre-treatments with tested ILs blended with10% (w/w) of water [58]. Different results were stated byBrandt et al. who pre-treated Miscanthus chips using[bmim][MeSO4] with 20.5% (w/w) of water [53]. In theseconditions the pre-treated biomass presented high di-gestibility. In contrast, the pre-treatment with dried[bmim][MeSO4] with water content below 0.3 wt% re-sulted in the formation of degraded biomass-IL compos-ite that was not enzymatically digestible. Therefore, acertain amount of water is necessary for successful pre-treatment of Miscanthus with [bmim][MeSO4]. However,after pre-treatment, [MeSO4] suffered from structuralchanges in the presence of water. It seems that water hy-drolyses the ester bonds in [MeSO4] to form MeOH and[HSO4] under the pre-treatment conditions applied. Toavoid the formation of a quaternary mixture of two ionicliquids ([bmim][MeSO4] and [bmim][HSO4]) plus twosolvents (water and methanol), [bmim][HSO4] with20.5% (w/w) of water as pre-treatment agent was chosen.After a 22-hour pre-treatment with this IL-aqueous mix-ture, most of lignin and hemicellulose was dissolved inthe liquid phase and 91% of the original glucan was re-covered in the resulted pulp. The authors also statedthat part of the dissolved hemicellulose undergoes hy-drolysis during pre-treatment. From the liquid phase,around 50% of the Klason lignin was recovered by pre-cipitation with water. Brandt et al. also tested pre-treatment with [emim][CH3COO] blending 20 vol% ofwater. The efficiency of this pre-treatment was similar tothat of the aqueous-[bmim][HSO4] solution, althoughthe sugar yields after enzymatic hydrolysis were found tobe lower [53].Wei et al. studied the pre-treatment of legume straw
with aqueous-[bmim][Cl] mixture [55]. This work wasdevoted to check the effect of temperature (50-170°C),time (0.5-4 hours), water content in the mixture (0-80%)and pressure (0.1-3.2 MPa) in the dissolution of legume
straw. At temperatures below 90°C almost no waterevaporation was observed due to possible strong interac-tions between water and [bmim][Cl]. For temperaturesabove 100°C the vaporised steam condenses and is re-cycled in the liquid phase. It was demonstrated that dis-solution was increased as temperature increases. Thevarious periods of time applied in the pre-treatment hadno effect on the dissolution. Pre-treatment performed athigh pressures, such as 3.2 MPa, increased the legumestraw dissolution while retaining the initial lignin con-tent found in the original biomass [55]. A 20% watercontent in the IL provided the best dissolution of legumestraw. These results enabled an optimised pre-treatmentat 150°C for 2 hours under ambient pressure, with amaximum of a 29.1 wt% legume straw dissolved in theaqueous-[bmim][Cl] mixture with water content of 20wt%. The optimised process demonstrated a lignin ex-traction of 48.2% from the original lignin content in thelegume straw. However, only 32.4% was recovered byadding water as a precipitate solvent. The biomass re-generation provided cellulose with an amorphous struc-tural conformation as occurred in pre-treatments withdried ILs [55]. The authors also evaluated the ability ofaqueous-[bmim][Cl] mixture in dissolving cotton pulpand no dissolution was observed. It was assumed that astrong interaction between cellulose and IL may existwith a proper amount of water to promote the dissol-ution of biomass [55].Triticale and wheat straws were also pre-treated by
aqueous-IL solutions and the results obtained were com-pared to pre-treatments using pure ILs [96,97]. Fu et al.observed a strong correlation between cellulose digest-ibility, lignin removal and crystallinity index of cellulosefor the pre-treatment of triticale straw in aqueous-[emim][CH3COO] solution [96]. Upon decreasing thewater content (95% to 0% w/w), an increased amount oflignin was extracted simultaneously with a crystallinereduction of the cellulose. However, after pre-treatmentperformances (150°C, 90 minutes) cellulose digesti-bility was substantially enhanced for aqueous-[emim][CH3COO] with 50% water content (w/w) achieving thehighest sugar yield (81.0%), higher than that observedfor pure [emim][CH3COO] (67.4%). Therefore, the pre-treatment efficiency is higher for pure IL in respect tolignin removal and crystallinity reduction. Nevertheless,if the goal of pre-treatment is to enhance the digestibilityof carbohydrates and maximise the release of fermenta-ble sugars, the pre-treatment with aqueous-[emim][CH3COO] was demonstrated to be more efficient, sinceit retains more carbohydrates and subsequently en-hances its digestibility [96]. Fu et al. provided a deeperstudy on the pre-treatment with aqueous-IL solutionsperforming an RSM statistical approach with wheatstraw and [emim][CH3COO] as the biomass and IL
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chosen [97]. They studied the effects of pre-treatmenttemperature (130-170°C), time (0.5-5.5 hour) and watercontent (0-100%) on lignin removal and cellulose/xylandigestibility. The optimised conditions were determinedto be 158°C, 3.6 hours with 50.5% water content (w/w)where 71.4% of the fermentable sugars recovery was de-termined as the predicted value. In comparison with thepre-treatment with pure [emim][CH3COO], the tempe-rature and time were found to also be beneficial for thedigestibility of carbohydrates from the pre-treatmentwith aqueous-[emim][CH3COO]. In this study, ligninwas largely extracted and the crystallinity reduced inpre-treatment with aqueous-[emim][CH3COO] leadingto a significant enhancement of the enzymatic hydroly-sis. However, it seems that a higher temperature andtime of pre-treatment are preferable to achieve thesame efficiency as pre-treatment with pure ILs [97].Nevertheless, the results for pre-treatment with aqueous-IL solution were similar to that observed in pure ILpre-treatment, but with less intensity. Due to thelower usage and easier recycling of ionic liquids, aswell as the reduced viscosity of the handling solution,pre-treatment with aqueous-IL mixture are potentiallycost-effective. However, a more envisaged investiga-tion is required, mainly about the acidic properties ofwater at high temperatures and its effect on biomasspre-treatment.
IL recycling and reuseThe reuse of recycled ILs for pre-treatment of biomassis a crucial factor for the economic efficiency of the pre-treatments with ILs, since ILs are more expensive thanconventional pre-treatment agents, such as ammoniaand sulphuric acid. Nevertheless, the IL recovery processhas been underexplored and for an effective operationmust be highly efficient.Generally, after the regeneration process, the liquid
stream is composed of an antisolvent with dissolved ILand soluble biomass compounds (lignin, soluble carbo-hydrates with low molecular weight, degradation prod-ucts, extractives and others) that were not precipitatedin the regeneration step. Depending on the IL recoveryprocess, these compounds can be recovered if furtherfractionation steps are applied. For instance, lignin couldbe precipitated by acidification of the liquid stream afterthe regeneration process [25,78]. Therefore, the IL re-covery is largely dependent on the antisolvent used, aswell as on the pre-treatment conditions applied thatguides to certain extend carbohydrates dissolution, ligninremoval and biomass degradation products present inthe liquid stream. Focused on maximising IL recovery,several processes were developed and further reusepre-treatments were performed to evaluate the pre-treatment efficiency after recycle [48,68,78,91,93]. Figure 4
represents three processes of IL recovery that were deve-loped up to present.Generally, the easiest way to recover the IL is to evap-
orate the antisolvent after the regeneration process, al-though this process would finish with several impuritiespresent in the IL, originated from the dissolved biomass[29,34,51,55,56,63]. For instance, [bmim][Cl] was recy-cled by simple evaporation of water from the aqueous/ILsolution obtained after separation of the regenerated leg-ume straw. The yield of the recycled [bmim][Cl] variedfrom 94% to 98%. However, a large quantity of impur-ities were determined, which changed the colourless ini-tial IL to an amber colour for the recovered IL [55]. Toovercome this limitation, sophisticated recovery pro-cesses were developed. Li et al. studied the [emim][DEP]recovery using two different antisolvents, namely waterand methanol [91]. The liquid stream after regenerationwas washed several times with diethyl ether. Duringwashings, salt was formed and then dissolved in chloro-form/methanol mixture. Subsequently, the remaining li-quid was passed through a column filled with neutralactivated alumina and the mixture solvent was thenevaporated to obtain the recovered [emim][DEP]. Theyield of the recycled IL using methanol as antisolventwas higher (96%) than that observed in the case of water(91%), although the amount of the regenerated biomasswas lower with methanol [91]. Tan et al. used [emim][ABS] for pre-treatment and NaOH aqueous solution asthe antisolvent, which resulted in a total mass of IL re-covery between 96.1% and 99.4% demonstrating a goodrecovery potential and high final purity [78]. Here the ILsolution was neutralised and water removed by evapor-ation resulting in collecting IL and NaCl as precipitatedsolids. Acetonitrile was added to dissolve selectively theIL, while NaCl remained as a solid to be filtrated in thenext step. The IL was finally recovered by acetonitrileevaporation [78]. Lan et al. also performed a similar ILrecovery process for [bmim][Cl] [93].Alternative for recovering ILs is provided by their abil-
ity to form a biphasic liquid-liquid system with theaddition of an aqueous solution containing a kosmo-tropic anion, such as phosphate, carbonate, or sulphate.Aqueous solutions of K3PO4 and K2HPO4 salts wereemployed resulting in a biphasic liquid-liquid systemcomprised by a salt-rich aqueous phase and an IL-richphase containing most of the lignin. The salting-out ef-fect leads to precipitation of a solid-phase rich in cellu-lose, simultaneously to the biphasic formation [48]. Shillet al. tested the IL recovery process with [emim][CH3COO] after pre-treatment of Miscanthus [48]. Theaqueous phosphate solution in the lower phase con-tained a high concentration of salt and trace amounts ofIL and sugars. The IL-rich phase was recycled after re-moval of water by evaporation achieving only a 72% IL
Figure 4 Schematic representation of the IL recovery technologies developed up to present. After pre-treatment the liquid stream iscomposed by IL, antisolvent and extracted material that can be recovered. The extracted material contains lignin, residual hemicellulose,degradation products and other soluble compounds. Generally, only lignin is recovered from liquid stream leading to losses of biomass residualmaterials at the end of the process. Depending on the IL recovery technology lower or higher pure IL can be obtained.
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recovery ready to be reused (without further solventevaporation). The part of the IL loss occurred in thewashing of the regenerated biomass [48]. Nevertheless,after the washing, water can be evaporated to recoverthe missing mass of the IL, although with noticeable im-purities (mostly hemicellulose and lignin). It was re-ferred that basicity of the K3PO4 solution presumablyresults in partial cleavage of lignin, allowing it to remainsoluble in the IL. The main advantage of this process isthe decreased amount of water present in the IL thatmust be evaporated and thus reduced energy costs.In an innovative process, Dibble et al. promoted phase
separation in the regeneration step by adding an an-tisolvent mixture constituted by ethanol and acetone inconcentration that creates a quaternary solution con-taining IL-water-ketone-alcohol [68]. The [emim][CH3COO] ionic liquid was selected as miscible withacetone by adjusting the quantities of water and ethanolin the solution. After regeneration, ethanol was main-tained in the solution with IL. Next, ethanol and part ofthe added acetone were distilled and dry acetone wasadded to the remained solution (mainly water and[emim][CH3COO]) creating a phase separation. Ali-phatic compounds were removed in the acetone phase
(the upper phase). After this process, several biomasscomponents remained dissolved in the IL solution com-prising low molecular weight carbohydrates, a portion ofthe original lignin, residual non-polar biomass compo-nents, and other compounds. The precipitation and re-covery of the majority of these residual solutes wereperformed by the addition of 2-propanol known as a lowpolarity solvent. This step reduced the accumulation ofthe very soluble biomass compounds in the IL. There-fore, after water distillation an efficient reuse of the ILfor a new pre-treatment could be attained [68]. After thepre-treatment process, the separation of dissolved lowmolecular weight carbohydrates, non-polar substances,low molecular weight lignin fragments, and other bio-mass components in ILs is a challenging task. In fact,the recycling process should remove these compoundsfrom the IL fraction as they are responsible for decreas-ing of the pre-treatment efficiency of the reused ILs. TheIL recovery process proposed by Dibble et al. [68] re-quires a minimal addition of reagents and it is assumedthat it would not result in significant IL degradation. Thereported recovery process resulted in 89% of the initial ILwithout the addition of acid or other catalysts for the re-covery of lignin and other solutes from the used IL [68].
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In the IL recovery process, the antisolvent is evapo-rated from the IL, thus volatile solvents could be eco-nomically desirable as they can reduce the energyrequired for distillation process comparing to waterdistillation. However, in biomass processing the use ofvolatile organic solvents, such as methanol, should beavoided from an environmental prospect. Thus, waterplays an important role as an antisolvent in the biomassregeneration. Therefore, an economical life-cycle assess-ment of the IL recovery should be evaluated.Apart from existent processes for IL recovery, a given
IL can be recycled if it is stable during the pre-treatmentand fractionation steps. Therefore, different pre-treatmentconditions should be considered for the optimal stabilityof the IL. In general, the studies that tested the IL recycle,made no mention of the degradation of the IL at the pre-treatment conditions applied and fairly good IL recoveryyields were obtained. One exception was observed in awork attempting to recycle [bmim][MeSO4] [53]. As ratedabove the [MeSO4] anion was partially hydrolysed to form[HSO4], promoted by the hydrolytic instability of the esterbonds in [MeSO4] anions under the conditions of the pre-treatment (120°C, 2–22 hours). In this case, mixtures ofthe ester and the hydrolysed form were produced [53].After IL recovery process there can still be several im-
purities present in the recovered IL. One of the majorimpurities is lignin that comes from the original biomassduring pre-treatment. Furthermore, lignin accumulationwas observed after reusing IL in a new pre-treatmentand this performance was continuously detected cycleafter cycle, without further purification of the IL[28,41,48,56,93]. Lee et al. concluded that lignin accu-mulation in [emim][CH3COO] occurs due to the highsolubility of lignin in this IL and that it does not affectthe efficiency of the pre-treatment of wood flour. More-over, they mentioned a similar pre-treatment efficiencyas that observed for fresh [emim][CH3COO] [28]. Infact, lignin accumulation and subsequently increasedconcentration in ILs can be advantageous for lignin re-covery as the facilitated precipitation of lignin could beaccomplished for further use in the generation of lignin-derived chemical products [28,41]. Notwithstanding, amaximum lignin removal from ILs is advisable becauseit may be necessary to prolong the lifetime of therecycled IL [48]. However, not only lignin can be presentin ILs after recycle, but also other contaminants, such asdegraded biomass, organic acids formed during pre-treatment or even introduced chemicals, water and othersolvents [93]. Lan et al. observed a clear increase incolour saturation after several reuse trials of [bmim][Cl]in the pre-treatment of sugarcane bagasse [93]. Ne-vertheless, 1H NMR analysis showed no obvious differ-ence between fresh and recycled IL. Change in colourintensity could indicate the existence of trace amounts
of contaminants in the recycled IL that were notdetected by 1H NMR analysis [93]. Tan et al. reportedthe loss of hemicellulose in the recovered IL reducingthe overall sugar recovery from the lignocellulosic biomassand making difficulties in the IL recovery process[71]. Hemicellulose and cellulose are described to bepartially degraded during pre-treatment with ILs. Afterseveral pre-treatment cycles in which the IL is reused,some degraded products, such as acetic acid seem toaccumulate in the recovered IL, and could catalysethe acidic fragmentation of carbohydrates in furtherreuse pre-treatments [56].Li et al. observed that reuse of [emim][CH3COO] in
wood pre-treatment did not affect the wood yield, al-though the composition of the wood was changed withthe number of IL recycles [56]. Therefore, the efficiencyof the recycled [emim][CH3COO] could be questionable.Haykir et al. determined the lignin extraction from cot-ton stalks with [emim][CH3COO] to be 47% via fresh ILand only a 27% for the first recycle of IL [63]. Wu et al.,by using the same IL in corn stover pre-treatment, re-ported a decrease in the extracted lignin content from44% in the first pre-treatment to a 20% in the tenth run,possibly due to lignin accumulation in the IL [41]. How-ever, no effect on subsequent fermentable sugar yieldswas demonstrated, thus [emim][CH3COO] could bereused up to 10 times, in spite of the lower extractionyield of lignin. The authors affirmed that β parameter of[emim][CH3COO] remains unaffected after each recyclebeside the accumulation of lignin. Since the interactionbetween the IL and lignin occurs mainly through thecation, the anions are still available to disrupt hydrogenbonds from the semi-crystalline cellulose. Therefore, nosignificant loss in the sugar yields of corn stover pre-treated with this recycled IL was observed [41]. Nguyenet al. tested a preceding treatment of rice straw with am-monia that provided an increase of the recycle numberof [emim][CH3COO] [51]. An insignificant increase inthe cellulose recovery from 82% to 83% was noticed forthe twentieth run with the recycled IL. The pre-treatment with the fifth time recycled [emim][CH3COO]showed a 97% glucose conversion in comparison withthe starting IL. Even for the tenth recycling cycle of IL,over 94% of the glucose conversion was attained [51].Cotton stalk samples pre-treated with recycled and fresh[emim][CH3COO] exhibited similar appearances pre-senting a lack of organised structure. In fact, accordingto a X-ray diffraction analysis, the crystallinity disruptionof cellulose to an amorphous form was unaffected after re-using [emim][CH3COO] [41,63]. The [emim][CH3COO]reuse was also feasible with respect to enzymatic di-gestibility, where the hydrolysis of pre-treated cotton stalkapparently showed no change after three times of IL re-cycling [41,63].
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The IL reuse was also investigated for other ILs[34,91,93]. The dissolution and regeneration rates ofpine wood obtained from both fresh and recycled[amim][Cl] were close to each other under comparablepre-treatment conditions. Moreover, the structure of thecellulose-rich regenerated material collected from therecycled IL was also similar to that obtained using fresh[amim][Cl], as verified by the 13C CP/MAS NMR, FTIR,XRD, and SEM techniques [34]. In the case of [bmim][Cl], the yield of regenerated sugarcane bagasse for therecycled IL decreased in comparison with that for thefresh IL [93]. The [emim][DEP] was reused in wheatstraw pre-treatment, for which it was noted that afterthe fifth run a 52.8% yield of reducing sugars wasobtained in comparison with 54.8% at the first pre-treatment. This result indicates that recycling and fur-ther reuse of [EMIM][DEP] did not rather affect thepre-treatment [91].
PerspectivesThe lignocellulosic biomass is a cheap and plentifulsource of energy, chemicals and materials present in thecomplex recalcitrant matrix. The current needs for non-fossil sources of energy and a strong development ofbiorefinery concept obligate to process biomass as anabundant feedstock. The classical methods of biomasspre-treatment are an interesting methodology to valorisebiomass; however, drawbacks of these methods limittheir wide application. That is why the emerging meth-odologies such as the application of ILs is a real alterna-tive. The numerous literature reports on this subjectprove these needs and demonstrate the enormous poten-tial of ILs. However, the complexity of both the biomassmatrix and ILs themselves makes the process very chal-lenging. That is why a large number of issues must to beconsidered in the designing of this kind of process. Un-fortunately, majority of literature reviews do not tacklemain problems, instead focusing only on some aspects.The number of potentially available ILs allows assumingthat “designer solvent – IL” can be found for each par-ticular problem. However, following the literature re-ports someone may have an impression that only acetateand chloride-based ionic liquids exist. Of course sometypes of ILs might be preferential for some particular ap-plication; nevertheless, this should not limit the study torediscover already existing ILs and their use in the bio-mass processing. It is especially weird as there arealready successful examples of IL use in the biomasscomponent processing with ILs characterised by favou-rable physico-chemical properties such as hydrophilicity/hydrophobicity, viscosity, melting point and others [98].Another important aspect related to the use of ILs inthe fractionation of biomass is the stability of ILs. ILsare considered to be generally extremely stable [10].
Although it must be understood that the IL thermal sta-bility measurements usually performed using differentialscanning calorimeter are mainly done for pure ILs in thecontinuously increasing temperature mode. So, it is clearthat these experiments do not represent the conditionsof the process with biomass. Pre-treatment of biomassin ILs is always a time consuming process that takesfrom minutes to dozen of hours and it takes place in thepresence of contaminants present in biomass, namelywater, acids and degradation products released fromhemicellulose hydrolysis upon pre-treatment. Defini-tively, under these much harsher conditions, the stabilityof ILs might be completely different, especially for ILscontaining a relatively strong base, such as [CH3COO](pKa = 4.77) [99] that show the slow, but noticeable de-composition after prolonged time [25,100].Pre-treatment of biomass allow producing cellulose,
hemicellulose and lignin and in some cases products oftheir hydrolysis. These fractions are the subject of fur-ther processing, very often with microorganisms (yeasts,bacterias, etc.), to produce ethanol or other high valueadded chemicals, e.g. xylitol. As it was aforementioned,ILs form an enormous class of compounds with widelytuneable properties including e.g. toxicity. Extensivestudies of the toxicity of ILs towards various organisms,including some of those used for biomass fraction pro-cessing exist [101]; however, additional systematic workon this topic is strongly demanded.Another factor that for now is rarely mentioned in the
literature; however, has a fundamental importance in therelation to the practical use of ILs is the economic andenergetic efficiency of the pre-treatment with ILs. As itwas shown in this review, the pre-treatment with ILs ismore effective and biomass fractions demonstrate higherpurity conversely to the fractions obtained from classicalpre-treatments. Therefore, in general, it can be statedthat the technological problems are overcome. However,the economic feasibility still might be questionable. Oneof reasons for this is an investment cost related to theprice of ILs. Nowadays, the availability and thus pricesare much more affordable; however, ILs are still moreexpensive than classical solvents. Nevertheless, it is im-portant to underline that IL prices at the industrial scaleare similar to organic solvents [102] and together withthe proven potential of IL reuse make processes moreeconomically feasible. Furthermore, with respect to theenergy consumption in the performance of the wholeprocess, IL use is also advantageous, as was alreadyshown by Yoon et al. [33] This pioneering work requiresmore efforts and needs to be continued to present thewhole picture and to guarantee a fair comparison withthe different methods available.The literature reports show that ILs are efficient tools
for biomass processing towards cellulose, hemicellulose
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and lignin fractions. The majority of the work dealingwith this subject focuses on the further processing ofcellulose that can be easily converted to cellulosicethanol widely used as biofuel. However, the two otherfractions, hemicellulose and lignin are more rarely con-sidered as important. The probable reason for this is thegreat diversity of both fractions that, on one hand,makes the process more difficult, but on the other opensthe room for a variety of commodities that can beobtained. The valorisation of these two diverse fractionsconstituted by different compounds depending on theraw material is especially important as it allows ob-taining products (e.g. xylitol, oligosaccharides, polyphe-nols, etc.) [103-105] with high commercial value thatcan make the whole process more economically feasible.The use of ILs in biomass pre-treatment is an import-
ant and interesting alternative; however, the aspectsdiscussed above are rarely tackled in the literature andrequire a profound and extensive analysis and study.Only after this careful and detailed analysis can therational assessment of biomass pre-treatment be per-formed. Unfortunately, the vast majority of papers pre-senting work about the pre-treatment of biomass withILs do not challenge these issues, thereby resulting inthe incomplete demonstration of the pros and cons ofbiomass processing with ILs.
ConclusionsLignocellulosic biomass is a widely abundant renewablefeedstock. The great diversity of lignocellulosic biomassopens a great opportunity for the production of variouscommodities using the wise valorisation processes. Ionicliquids have proven the inordinate potential in reactionsand separation including biomass processing. This re-view shows that ILs have potential to be used in the bio-mass pre-treatment. As it was demonstrated, ILs can besuccessfully used to obtain cellulose, hemicellulose andlignin fractions with purity and efficiency equal or super-ior to the classical pre-treatment methods. The exploit-ation of ILs in the lignocellulosic biomass processingdemonstrates the enormous potential in this researcharea, especially in the context of a biorefinery; however,an extensive research is still required to better under-stand and predict outcomes of these processes.
Competing interestsThe authors declare that they have no competing interests.
Authors’ contributionsAMCL and ARCM wrote the principle part of the review. KJG did a literaturedata collection. EBL was responsible for the compilations of the literaturedata in the tables. RBL was responsible for the overall work with theparticular attention to perspectives and conclusions. All authors read andapproved the final manuscript.
AcknowledgmentsThis work was supported by the Fundação para a Ciência e a Tecnologia(FCT, Portugal) through grants SFRH/BPD/26356/2006, PEst-C/EQB/LA0006/2011 and Programme Ciência 2008, and by the European Commission for thefinancial support of the PROETHANOL2G Project (FP7-ENERGY-2009-BRAZIL;Grant agreement: 251151). Authors wish to acknowledge LNEG for thefinancing of the ERA-IB project (Products from lignocelluloses-EIB.10.013).
Author details1Laboratório Nacional de Energia e Geologia, Unidade de Bioenergia,1649-038 Lisbon, Portugal. 2Departamento de Química, Faculdade deCiências e Tecnologia, REQUIMTE, Universidade Nova de Lisboa, 2829-516Caparica, Portugal.
Received: 31 January 2013 Accepted: 19 April 2013Published: 22 May 2013
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doi:10.1186/2043-7129-1-3Cite this article as: da Costa Lopes et al.: Ionic liquids as a tool forlignocellulosic biomass fractionation. Sustainable Chemical Processes 20131:3.
