Strong hydrogen-bonding basicity ф-value in KAT values) is now recognized as the most important property of ILs with high cellulose dissolution. Viscosity of ILs is the second key factor causing cellulose dissolution at low temperature conditions. However, a rational design of ILs with a dissolution property of cellulose has not yet been established. We discuss in this chapter how to accomplish the design of ILs with high cellulose dissolution from the standpoint of nature.

Remsing et al. [23] reported based on their 13C and 35/37Cl NMR studies that there was a stoichiometric interaction between the chloride anion and the cellulose hydroxyl groups, and this might be the key driving force of cellulose dissolution in this IL (Fig. 4.3) [23].

Inspired by their result, we hypothesized that enhanced interaction of a certain anion or cation of ILs between hydroxyl groups in the cellulose molecule might be the key factor causing cellulose dissolution and we might be able to obtain a hint on how to design such anion or cation from nature. Focusing on the structure of hydrolyzing enzyme of cellulose (cellulase), we found that amino acid ILs were strongly capable of dissolving cellulose: N, N-diethyl, N-methyl, N-(2-methoxy) ethylammonium alanate ([N221(ME)][Ala]) worked as an excellent solvent for cel­lulose dissolution among ILs whose anion part was natural amino acid [24].

Hydrolysis of solid cellulose is achieved by cellulases such as endoglucanase (EGs) and cellobiohydrolases (CBHs) [25, 26]. The former can hydrolyze internal p-1,4-glycoside bonds in a cellulose polymer in the amorphous regions within the cellulose micro-fibril, and the latter can act on the free ends of cellulose polymer chains. Both types of cellulases have cellulose-binding modules that facilitate their

adsorption onto crystalline cellulose, bringing the catalytic domains physically close to their site of action (Fig. 4.4) [25, 26]. We looked at what the protein sequences of several cellulases were causing particularly in the area of substrate­binding cleft, and recognized that glucosyl-binding sites of cellulases were fre­quently formed by the exposed surface of aromatic side-chains of protein residues [25, 26]. Three of the four binding sites making up the enclosed cellulose-binding tunnel reportedly contain the tryptophan (W), asparagine (N), and isoleucine

(I) residue side-chains for Trichoderma reesei Cel6A (CBH II) [25, 26]. Therefore, it was expected that ILs made from amino acids might have an affinity toward a certain part of cellulose.

Ohno and co-workers prepared ILs that contained amino acid moieties as anion parts [21,27]. Since the hydrogen-bonding basicity of amino acid salts was reported to be high [27], amino acid ionic liquids are expected to display good cellulose dissolving ability.

Based on these results, the dissolving property of 1-butyl-3-methylimidazolium tryptophan ([C4mim][Trp]) against cellulose was tested using microcrystalline
cellulose (Avicel®) as a model compound. However, the cellulose did not dissolve at all in this IL. Further evaluation of tryptophan salts with ammonium, phospho — nium, or pyrridinium cation, revealed that choice of cation was also a key point in designing an IL with cellulose dissolution capability: N-(2-methoxyethyl),N, N — diethyl, N-methylammonium tryptophan ([N221(ME)][Trp]) dissolved cellulose (5 wt% vs. IL) at 100 °C [24]. Encouraged by the results, we prepared [N221(ME)] salts with natural amino acids and carefully evaluated their cellulose dissolution properties against the model cellulose (Avicel) (Table 4.2). Among 20 types of amino acid salts, we found that [N221(ME)][Ala] worked best to dissolve cellulose with 12 wt% versus solvent: the second solvent most effective was lysine salt ([N221 (ME)][Lys]) (11 wt%) and the third was ornitin salt ([N221(ME)][Orn]). Threonine ([N221(ME)][Thr]) and isoleucine ([N221(ME)][Ile]) salts also showed similar solubil­ity against the cellulose (7 wt%) [24]. We expected that amino acid might have affinity with a certain part of cellulose and cause its dissolution in the amino acid ionic liquid. The results reached our expectations though the details were slightly different; one of these we had anticipated, however, because there was no alanine residue near the entrance part of the cellulases [25, 26].

Many amino acid ILs dissolved cellulose, except for glutamic acid salt and all amino acids had high p-values [27]. Hence, we fixed the anionic part to alanin, and the cationic portion was re-evaluated (Fig. 4.5). It was thus confirmed that cellulose solubility was strongly dependent on the cationic part. High cellulose solubility was recorded for N, N-bis(2-methoxyethyl),N-ethyl-N-ethylammonium ([N22(ME)2]), N, N, N-tris(2-methoxyethyl),N-ethylammonium ([N2(ME)3]), N-(2-thiomethoxyethyl), N, N-diethyl, N-methylammonium ([N221(MTE)]), N-methyl-N-ethoxyethylpyr-

rolidium ([P1(ME)]) salts, in a range of 12 to 11 wt%. On the contrary, no dissolution of cellulose took place in [P444ME][Ala] or [PyME][Ala] salt. Interestingly, the presence of the methoxyethyl group on the ammonium cationic part strongly modified cellulose solubility: better dissolution was obtained for [N221ME][Ala], while poor solubility was obtained for N-butyl-N, N-diethyl, N-methylammonium alanine([N4221][Ala]). However, both the methoxyethoxymethyl substituted salt ([N221(MEM)]) and N, N,N, N-tetra(methoxyethyl)ammonium ([N(ME)4]) alanine showed poor cellulose solubility (Fig. 4.5). These results clearly indicate that cellulose solubility is determined not only by the physical characteristics of the solvent shown as KAT values, but also by the affinity of a certain group interaction of ionic liquids with cellulose might be an important factor of cellulose dissolution in the ILs.

We next investigated cellulose solubility against various ILs which have [N221 (ME)] as a cationic part. Although 10 wt% of cellulose dissolves in [N221(ME)]Cl, this salt requires a higher temperature (over 120 °C) and a longer mixing time over [N221(ME)][Ala]. Slight decomposition of IL was observed under the conditions used [24]. No dissolution of cellulose was observed when [N221(ME)]Br, hexafluor- ophosphate (PF6), N, N-bis(trifluoromethyl)sulfonylamide (NTf2), 2,2,3,3,4,4,5,5- octafluoropentyl sulfate (C5H8), or 2-aminoethylsulfonate (taurine), was used as solvent. [N221(ME)] salts with hydrogen oxide also showed poor dissolution prop­erties [24]. Poor solubility was also recorded for N, N-dimethylalaine or

N-Boc-alanine salts compared to the alanine salt (Fig. 4.6). The presence of amino group might play an important role in dissolving cellulose [24]. Therefore, we anticipate that the amino group of [Ala] may interact with a certain part of cellulose and contribute to breaking its hydrogen bond network. Liquid ammonia reportedly changes the crystalline phase of naturally occurring cellulose and dissolves it by allowing the ammonia molecules to penetrate the cellulose fibril [28, 29]. From these results, we assume that the amino group interposed the hydrogen bonding between the cellulose and caused dissolution of cellulose in amino acid ILs. However, since the cellulose solubility is also modified by the cationic part of the IL, cation might play an important cooperative role in the mechanism for cellulose dissolution, although its origin is still unclear.

Addition of an anti-solvent like water or ethanol to the cellulose/IL solution causes precipitation of the dissolved cellulose and the structure of the regenerated cellulose changes to a disordered form. Pretreatment of cellulose increases the surface area accessible to water and cellulases are believed to improve the hydro­lysis rate [30, 31]. Therefore, many researchers have attempted to hydrolyze regenerated cellulose in order to improve the hydrolysis rate by a cellulase; the dissolution of microcrystalline cellulose with [C4mim]Cl and the rapid
precipitation with water induced an increase of the amorphous region in the regenerated cellulose and enhanced the initial enzymatic hydrolysis rate [30, 31].

It was reported that cellulose regenerated from ionic liquid solution showed Type II crystalline form [31]. In fact, we confirmed that the regenerated cellulose from [N221(ME>][Ala] solution had only Type II form [24]. The X-ray diffraction patterns of the microcrystalline cellulose film (Avicel®) and that of the regenerated one were compared: the regenerated cellulose exhibited the typical diffraction patterns of Type II cellulose at 20 = 20.16° and 21.76° [32]. The results indicate that the transformation from Type I to Type II occurred after the dissolution and regeneration in [N221(ME)][Ala] [24].

Interestingly, we further found that crystal form of the regenerated cellulose was dependent on the dissolution solvent [24, 33]: 7 wt% of cellulose was dissolved in [N221(ME)][(MeO)(H)PO2] and the regenerated cellulose was a mixture of Type I and II.

Mizuno et al. [33] recently reported that the disordered chain region was increased in the order of [N221(ME)][Ala] < [C2mim][OAc] < [C2mim] [(EtO)2PO2] < [C2mim]Cl [33]: regenerated cellulose treated with [C2mim]Cl contained larger amorphous regions than the others. On the contrary, that of [N221 (ME)][Ala] had a larger amount of cellulose II crystalline structure and less suscep­tibility to enzymatic degradation than others; this suggests that the enzymatic hydrolysis rate of regenerated cellulose should increase by the same order. On the other hand, the order of degree of polymerization of the cellulose was [N221(ME)] [Ala] > [C2mim][OAc] > [C2mim][(EtO)2PO2] > [C2mim]Cl; treatment with [N221(ME)][Ala] is therefore much more suitable to use in preparing regenerated cellulose fiber.