Ionic liquids enzymatic reactions

Irimescu R, Kato K (2004) Investigation of ionic liquids as reaction media for enzymatic enantioselective acylation of amines. J Mol Catal B Enzym 30 189-194... 

Ionic Liquids as Reaction Media in Enzymatic Synthesis... 

Enzymatic reactions are often performed in aqueous buffer solution addition of increasing amounts of ionic liquids sometimes caused precipitates of unknown composition. 

To maintain enzymatic activity a minimal amount of water has to be present, best described by the water activity. However, water present in the reaction system may cause hydrolysis of some ionic liquids. 

In the first publication describing the preparative use of an enzymatic reaction in ionic liquids, Erbeldinger et al. reported the use of the protease thermolysin for the synthesis of the dipeptide Z-aspartame (Entry 6) [34]. The reaction rates were comparable to those found in conventional organic solvents such as ethyl acetate. Additionally, the enzyme stability was increased in the ionic liquid. The ionic liquid was recycled several times after the removal of non-converted substrates by extraction with water and product precipitation. Recycling of the enzyme has not been reported. It should be noted, however, that according to the log P concept described in the previous section, ethyl acetate - with a value of 0.68 - may interfere with the pro-... 

To carry out the enzymatic amidation of carboxylic acids, normally two strategies are considered the use of ionic liquids or the removal of water from the reaction media at high temperature or reduced pressure. For instance, one of the first examples of the use of ionic liquids in biocatalysis has been the preparation of octanamide from octanoic acid as starting material and ammonia in the presence of CALB (Scheme 7.3) [11]. 

Irimescu and Kato have recently described an interesting example of enzymatic KR in ionic liquids instead of organic solvents (Scheme 7.4) [12]. The resolution with CALB is based on the fact that the reaction equilibrium was shifted toward the amide synthesis by the removal of water under reduced pressure. Nonsolvent systems have been also employed in this enantioselective amidation processes, reacting racemic amines with aliphatic acids. The best reaction conditions for the conversion of acids to amides was observed using CALB at 90 °C under vacuum. Meanwhile, no... 

S., Kula, M.R., and Kragl, U. (2003) Enzymatic Condensation Reactions in ionic liquids, in Ionic Liquids as Green Solvents, ACS Symposium Series, Vol. 856 (eds R.D. Rogers and... 

The use of ionic liquids (ILs) to replace organic or aqueous solvents in biocatalysis processes has recently gained much attention and great progress has been accomplished in this area lipase-catalyzed reactions in an IL solvent system have now been established and several examples of biotransformation in this novel reaction medium have also been reported. Recent developments in the application of ILs as solvents in enzymatic reactions are reviewed. 

Figure 1 The first enzymatic reaction conducted in a pure ionic liquid solvent...

In a recent review, some positive attributes of ionic liquids in biocatalysis were discussed 273). An example was given, which compares the enzymatic performance of Pseudomonas cepacia lipase (PCL)-catalyzed reactions as a function of the solvent polarity in both organic and ionic solvents, as shown in Fig. 17. The PCL shows no activity in organic solvents in the polarity range of the ionic liquids, but it is active in the ionic liquids. 

The beneficial effect of the hydrophobicity of [BMIM]PFg was shown to extend to other enzymes a remarkably enhanced enantioselectivity was observed for lipases AK and Pseudomonas fluorescens for the kinetic resolution of racemic P-chiral hydroxymethanephosphinates (Scheme 31) (278). The ee values of the recovered alcohols and the acetates were about 80% when the enzymatic reactions were conducted in the hydrophobic [BMIMJPFg. In contrast, there was little enantioselectivity (<5%) observed with the enzymes in hydrophilic [BMIM]BF4. The lack of stereoselectivity in [BMIM]BF4 was attributed to the high miscibility of [BMIM]BF4 with water. The relatively hydrophilic ionic liquid is capable of stripping off the essential water from the enzyme surface, leading to insufficient hydration of the enzyme and a consequently strong influence on its performance (279). 

In the synthesis of A-acetyllactosamin from lactose and A-acetylglucosamine with (3-galactosidase (289,290), the addition of 25 vol% of the water-miscible ionic liquid [MMIM][MeS04] to an aqueous system was found to effectively suppress the side reaction of secondary hydrolysis of the desired product. As a result, the product yield was increased from 30 to 60%. Product separation was improved, and the reuse of the enzymatic catalyst became possible. A kinetics investigation showed that the enzyme activity was not influenced by the presence of the ionic liquids. The enzyme was stable under the conditions employed, allowing its repeated use after filtration with a commercially available ultrafiltration membrane. 

The enzymatic enantioselective hydrolysis of esters of naproxen and ibuprofen has attracted considerable attention because the (S)-enantiomers of these nonsteroidal anti-inflammatory drugs (NSAIDs) are the pharmacologically active isomers. These reactions have been successfully performed in a range of ionic liquids (Figure 10.10) [60, 65, 121]. 

Proteases have been much less studied than lipases in ionic liquid media and generally require the presence of water for activity. We note that the thermolysin-catalyzed amide coupling of benzoxycarbonyl-L-aspartate and L-phenylalanine methyl ester into Z-aspartame in [BMIm][PF6] was an early example of an enzymatic reaction in an ionic liquid medium [8]. 

In biphasic reactors or two-phase partitioning bioreactors (TPPB), the substrate is located mostly in the immiscible phase and diffuses to the aqueous phase. The enzyme catalyzes conversion of the substrate at the interface and/or in the aqueous phase. The product/s of the reaction then may partition to the organic phase. The system is self-regulated, as the substrate delivery to the aqueous phase is only directed by the partitioning ratio between the two phases and the enzymatic reaction rate [53]. The use of ionic liquid/supercritical carbon dioxide for enzyme-catalyzed transformation is gaining attention [69]. 

Reactions proceed faster and more smoothly when the reactants are dissolved, because of diffusion. Although reactions in the solid state are known [1] they are often condensations in which a molecule of water is formed and reaction takes place in a thin film of water at the boundary of the two solid surfaces. Other examples include the formation of a liquid product from two solids, e.g. dimethylimidazolium chloride reacts with aluminum chloride to produce the ionic liquid, dimethylimidazolium tetrachloroaluminate [2]. It is worth noting, however, that not all of the reactant(s) have to be dissolved and reactions can often be readily performed with suspensions. Indeed, so-called sol-id-to-solid conversions, whereby a reactant is suspended in a solvent and the product precipitates, replacing the reactant, have become popular in enzymatic transformations [3]. In some cases, the solvent may be an excess of one of the reactants. In this case the reaction is often referred to as a solvolysis, or, when the reactant is water, hydrolysis. 

Ionic liquids were initially developed as solvents for electrochemical applications. The electrochemical window of clean ionic liquids can be huge/11 allowing for a wide range of redox reactions/2,31 It has further been demonstrated that they are also suitable solvents for enzymatic oxidations14 71 but both topics are beyond the scope of this book. Only transformations that involve the metal-catalysed addition of oxygen to unsaturated carbon bonds as well as the oxidation of alcohols, aldehydes and ketones to their corresponding ketones, carboxylic acids and esters shall be discussed in this chapter. 

Although the use of enzymes in ionic liquids has been explored, the enzymatic carbon-carbon bond forming the reactions in ionic liquids has not been studied in detail. Since aldol reactions catalyzed by the aldolase antibody 38C2 in buffer... 

Several thermodynamic and kinetic behaviors of enzyme-catalyzed reactions performed in ILs, with respect to enzymatic reactions carried out in conventional solvents, could lead to an improvement in the process performance [34—37]. ILs showed an over-stabilization effect on biocatalysts [38] on the basis of the double role played by these neoteric solvents ILs could provide an adequate microenvironment for the catalytic action of the enzyme (mass transfer phenomena and active catalytic conformation) and if they act as a solvent, ILs may be regarded as liquid immobilization supports, since multipoint enzyme-1L interactions (hydrogen. Van der Waals, ionic, etc.) may occur, resulting in a flexible supramolecular not able to maintain the active protein conformation [39]. Their polar and non-coordinating properties hold considerable potential for enantioselective reactions since profound effects on reactivities and selectivities are expected [40]. In recent years attention has been focused on the appUcation of ILs as reaction media for enantioselective processes [41—43]. 

As can be seen, immobilized CALB efficiently catalyzes the acylation of aU polyhydroxylated compounds in the ionic liquid media used, leading to high conversion yields. The reaction rates for the enzymatic acylations are summarized in Table 9.3. Higher reaction rates were obtained for aU polyhydroxylated compounds tested when [bmim]BF4 was used as the reaction medium. It is interesting to note that the solubilities of esculin, salicin, helicin, naringin, and silybin at 60 C were approximately 68, 23,40,100, and 82 mM, respectively, in [bmim]BF4. On the other hand, in [bmim]PFs and acetone, in which the reactions rates were lower, the solubilities of the aforenamed compounds were 20, 5.5,19.5,1.5, and 7.2 mM and 6.5,... 

The monoacylated derivative (6"-0-butyrate) was detected as the major product by H PLC analysis for all enzymatic reactions, while a diacylated derivative was also detected and confirmed by MS analysis [17]. As it can be seen in Table 9.3, higher regioselectivity of the process was observed in [bmim]BF4 than in [bmim]PF,5, while the selectivity was even lower in acetone. The lower regioselectivity of the enzymatic acylation of polyhydroxylated compounds in [bmim]PF,5 as well as in the organic solvent could be related to the lower solubility of unmodified phenolic substrates in these media, compared to that of their monoacylated derivatives [5, 16, 17]. On the other hand, the enhanced solubility of phenoHc substrates in [bmim]BF4 could explain the increased regioselectivity observed in this ionic liquid. 

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