Due to their unique proprieties, ILs have been used for biodiesel production processes through lipase catalyzed transesterification (alcoholysis) of vegetable oils (or animal fats) (Fig. 11.2) by several research groups [27—34]. Biodiesel is a renewable diesel fuel that is also known as FAME (fatty-acid methyl esters).

The enzymatic transesterification method offers many advantages over the chemical methods, such as mild reaction conditions, low energy demand, low waste treatment, the reusability of enzymes (lipases in most cases), flexibility in choosing different enzymes for different substrates, and the fact that it allows a small amount of water to be present in substrates. Besides, in chemical processes, some oil or fats may need a pretreatment for deacidification, depending on the composition of the materials, to remove free fatty acids (FFAs), which form soap with alcohols. Lipase can convert both triglycerides and FFAs into biodiesel [2]. Some authors have even successfully used waste cooking oil to obtain enzy­matic biodiesel which may be a promising alternative for reducing the cost of biodiesel [27, 35—38].

The production yield was improved markedly when immobilized Candida antarctica lipase B (CALB) on an acrylic resin was used as a biocatalyst compared with other microbial lipases [27, 29, 30, 39, 40]. Ha et al. [30] screened 23 ionic liquids as solvents in the production of biodiesel from soybean oil using Candida antarctica lipase as catalyst. [EMIM][TfO] produced the highest biodiesel yield (80 % in 12 h of reaction), a yield that was better than for the solvent-free system and other commonly used solvents (tert-butanol). Nineteen ILs were studied to determine their effectiveness as solvents in the transesterification process using Burkholderia cepacia lipase (BSL) as catalyst [32]. The ionic liquid used have combinations of cations and anions, being the cations based mainly on imidazolium, while the anions were [NTf2] and [PF6] to get a suitable reaction media (Table 11.1).

Lipase-catalyzed methanolysis when conducted in a solvent-free medium led to the deactivation of lipase with increased molar ratio of methanol to sunflower oil >3 [45]. A similar deactivation of lipases was also observed during lipase cata­lyzed methanolysis in a biphasic oil-aqueous system for FAME production [46]. Methanolysis was conducted at different molar ratios of methanol to oil

(4:1 to 10:1) and similar results were obtained. Thus the concentration of methanol did not have a great effect on product formation in the presence of IL, which protects the lipase from methanol-induced deactivation.

Among the various types of ILs used, hydrophobic ILs were found to be the most effective for the production of biodiesel, the biodiesel yield increasing with both cation chain length and IL hydrophobicity, and decreasing when ILs with strong water miscible properties were used ([BMIM][NTf2], [EMIM][PF6], AMOENG 100, AMOENG 102, [C16MIM][NTf2] or [C18MIM][NTf2] etc). Hydrophilic ILs were not suitable as solvent in enzyme-catalyzed transesterification as only 10 %

FAME yield was obtained for [HMIM][BF4], while no FAME was observed when [BMIM][BF4] was used as the solvent [33].

This trend can be explained in terms of methanol-induced enzyme deactivation. Hydrophobic ILs protects the enzyme from such deactivation because lipase is entrapped in the IL matrix. The most notable advantages of the use of ILs in such bioconversions are that the biodiesel can be separated by simple decantation and the recovered ionic liquid/enzyme catalytic system can be re-used several times with­out loss of catalytic activity and selectivity. More recently, our group [27] used [C16MIM][NTf2] as a homogeneous reaction mixture and, when the reaction was complete, a triphasic system was created through the appearance of a FAMEs phase (upper layer), a glycerol phase (middle layer) and a lower layer with the ionic liquid containing the enzyme, which could be solidified by decreasing the reaction temperature of the media (the melting point for [C16MIM][NTf2] is 42.6 °C), in this way facilitating extraction of the biodiesel product. Furthermore, ILs provides the ideal medium for removal of the by-product glycerol, thus accounting for the increase in biodiesel yield. A promising strategy employed by Lai et al. [47] was to use cross-linked enzyme aggregates (CLEAs) of lipase from Penicillium expansum as catalyst for biodiesel production in [BMIM][PF6] from microalgal oil, with a conversion of 85.7 % in 48 h.