This chapter discusses selected recent applications of organic carbonates as reactive solvents in biorefineries and biotechnology, with emphasis on the diversity of options and functions that these solvents may play: mere solvents, reagents or extractive media. It must be noted that organic carbonates have been traditionally synthesized using activated carbonylic derivatives such as phosgene, or employing synthesis‐gas effluents (containing CO). Combined with efforts to introduce more sustainable syntheses for organic carbonates, it is worth investigating whether the solvent capabilities and reactivities of such solvents would fit within several biorefinery‐based processes. organic carbonates exert a potential double profile: on the one hand, they may be employed as solvents; on the other, their intrinsic reactivity may be used to trigger processes (e.g. acting as reagents while dissolving other substrates). Within biocatalysis, this has been the case of dimethyl carbonate (DMC), which has attracted considerable interest as solvent and as reagent to produce, for instance, glycerol carbonate. Bio‐Based Solvents Bio‐Based Solvents  Related  Information Details © 2017 John Wiley & Sons, Ltd. Keywords biocatalysis biorefineries biotechnology dimethyl carbonate enzymatic synthesis non‐conventional media organic carbonates reactive solvents Publication History @en

Arabidopsis thaliana hydroxynitrile lyase (AtHNL) catalyzes the selective synthesis of (R)-cyanohydrins. This enzyme is unstable under acidic conditions, therefore its immobilization is necessary for the synthesis of enantiopure cyanohydrins. EziG Opal is a controlled porosity glass material for the immobilization of His-tagged enzymes. The immobilization of His6-tagged AtHNL on EziG Opal was optimized for higher enzyme stability and tested for the synthesis of (R)-mandelonitrile in batch and continuous flow systems. AtHNL-EziG Opal achieved 95% of conversion after 30 min of reaction time in batch and it was recycled up to eight times with a final conversion of 80% and excellent enantioselectivity. The EziG Opal carrier catalyzed the racemic background reaction; however, the high enantioselectivity observed in the recycling study demonstrated that this was efficiently suppressed by using citrate/phosphate buffer saturated methyl-tert-butylether (MTBE) pH 5 as reaction medium. The continuous flow system achieved 96% of conversion and excellent enantioselectivity at 0.1 mL min−1 . Lower conversion and enantioselectivity were observed at higher flow rates. The specific rate of AtHNL-EziG Opal in flow was 0.26 mol h−1 genzyme−1 at 0.1 mL min−1 and 96% of conversion whereas in batch, the immobilized enzyme displayed a specific rate of 0.51 mol h−1 genzyme−1 after 30 min of reaction time at a similar level of conversion. However, in terms of productivity the continuous flow system proved to be almost four times more productive than the batch approach, displaying a space-time-yield (STY) of 690 molproduct h−1 L−1 genzyme−1 compared to 187 molproduct h−1 L−1 genzyme−1 achieved with the batch system.@en

The question of how to distinguish between lipases and esterases is about as old as the definition of the subclassification is. Many different criteria have been proposed to this end, all indicative but not decisive. Here, the activity of lipases in dry organic solvents as a criterion is probed on a minimal α/β hydrolase fold enzyme, the Bacillus subtilis lipase A (BSLA), and compared to Candida antarctica lipase B (CALB), a proven lipase. Both hydrolases show activity in dry solvents and this proves BSLA to be a lipase. Overall, this demonstrates the value of this additional parameter to distinguish between lipases and esterases. Lipases tend to be active in dry organic solvents, while esterases are not active under these circumstances.@en

The first enantioselective synthesis was the selective addition of cyanide to benzaldehyde catalysed by a hydroxynitrile lyase (HNL). Since then these enzymes have developed into a reliable tool in organic synthesis. HNLs to prepare either the (R)- or the (S)-enantiomer of the desired cyanohydrin are available and a wide variety of reaction conditions can be applied. As a result of this, numerous applications of these enzymes in organic synthesis have been described. Here the examples of the last decade are summarised, the enzyme catalysed step is discussed and the follow-up chemistry is shown. This proves HNLs to be part of main stream organic synthesis. Additionally the newest approaches via immobilisation and reaction engineering are introduced.@en

The hydroxynitrile lyase from Prunus amygdalus was immobilized on Celite R-633. The immobilized enzyme could successfully be utilized in buffer saturated MTBE and excellent conversions of benzaldehyde to R-mandelonitrile were observed. No leaching occurred. To achieve high enantioselectivities, the suppression of the undesired background reaction was essential. This could be achieved by high enzyme loadings and the tight packing of the immobilized enzymes. When the immobilized enzyme is loosely packed, both the enzyme catalysis and the background reaction accelerates and only a modest enantioselectivity is observed. The enzyme was recycled for up to ten times, with some loss of activity and also enantioselectivity after 5 cycles, independent of packing.@en

The enzymatic oxidation of amino alcohols was studied to address the long-standing problem of product stability. Amino aldehydes, highly sought and unstable compounds, can be generated under mild conditions if they are immediately protected. Utilizing a range of alcohol dehydrogenases (ADHs) and semicarbazide as a scavenger, the enantioselective synthesis of protected amino aldehydes is possible. Glycerol dehydrogenase from Gluconobacter oxydans (GoGDH) displayed excellent enantioselectivity but limited substrate scope, whereas horse liver ADH catalyzed a broad range of conversions with low enantioselectivities.@en

Enzymes are supreme catalysts when it comes to high enantiopurities and their immobilization will pave the way for continuous operation. In this context, we show the covalent immobilization of hydroxynitrile lyases HbHNL (from Hevea brasiliensis) and MeHNL (from Manihot esculenta) in a siliceous monolithic microreactor for continuous operation. A thorough characterization of the immobilized HNLs on mesoporous silicates indicated the conditions essential for a successful immobilization. Their application in a continuous flow system enabled a remarkably fast (3.2 min) production of chiral cyanohydrins with high conversion (97%) and high ee (98%) using minimal enzyme loading (STY = 71 g L -1 h -1 mg protein -1 ). MeHNL showed increased operational stability, possibly due to a structural difference. The continuous flow microreactor outperformed batch systems, demonstrating the advantage of the mesoporous/macroporous environment for the expression of enzyme activity and the favorable characteristics of the microreactor. Overall, the system shows great potential for future industrial application of biocatalytic asymmetric syntheses. @en