Microwave-Assisted Kinetic Resolution of Homochiral (Z)-Cyclooct-5-ene-1,2-diol and (Z)-2-Acetoxycyclooct-4-enyl Acetate Using Lipases

Over the last decade, the use of biocatalysts has become an attractive alternative to conventional chemical methods, especially for organic synthesis, due to their unusual properties. Among these enzymes, lipases are the most widely used, because they are cheap, easily available, cofactor-free, and have broad substrate specificity. Combined to microwave heating in non-aqueous medium, recent results suggest that irradiation may influence the enzyme activity. This Communication reports the benefits of lipases and the microwave irradiation on the kinetic resolution of racemic homochiral (Z)-cyclooct-5-ene-1,2-diol and (Z)-2-acetoxycyclooct-4-enyl acetate. In order to best achieve the kinetic resolution, different parameters were studied including the type of lipase, the temperature, the impact of microwave power compared to conventional heating. Optimization of the reaction parameters lead to the obtainment of highly enriched or enantiopure diols and diesters in a clean, efficient and safe way.


Introduction
Enantiomerically pure vicinal diols are versatile chemical scaffolds for the production of flavors and fragances. As part of our work on the enantioselective synthesis of methyl jasmonate derivatives from optically active bicyclo [3.3.0]octane derivatives by transannular cyclization, we first needed to prepare enantiopure homochiral (1R,2R)-and (1S,2S)-5-cyclooctene-1,2-diols. In recent years, the development of biocatalysts for organic synthesis has become an attractive alternative to conventional chemical methods. Among those biocatalysts, lipases have become very popular in both academic and industrial sectors because they are inexpensive, easily available, cofactor free and have a broad substrate specificity [1]. We decided to focus our interest on microwave-assisted lipase-mediated kinetic resolution involving CaLB (lipase B from Candida antartica) or PS (Pseudomonas cepacia)-catalyzed acetylation of diol. The use of microwave irradiation in biocatalysis can enhance the enzyme activity, for example in resolution reaction, in specific oxido-reduction reaction or hydrolysis. Combined to non-aqueous medium, recent results suggest that microwave irradiation can have also influence the enzyme stability and activity, in addition to altering/enhancing reaction rates and/or enantioselectivities, called non-thermal microwave effects [2][3][4][5][6][7][8]. However, the exact role of microwave irradiation on enzymes still remains unresolved [9]. To better comprehend the influence of microwave irradiation on a biocatalyst, we decided to compare the lipase-catalyzed resolution of difunctionalized compounds, under conventional and microwave irradiation heating. We report herein our studies on microwave assisted lipase resolution of homochiral (1R,2R)-and (1S,2S)-5-cyclooctene-1,2-diols 2 and their diesters 4, by varying the irradiation power and reaction temperature.

Results and Discussion
In the context of the growing general interest for reducing energy costs, heating chemical reactions under microwave irradiation is a useful approach for achieving higher reaction kinetics and synthesizing cleaner products [10][11][12][13]. In combination with microwave technology and lipases, we wished to examine the synthesis of chiral cyclooctenic diols and diesters starting from cycloocta-1,5-diene using microwave technology. rac-Diols 2 and rac-diacetates 4 were initially prepared from cycloocta-1,5-diene in a three-steps sequence including epoxidation, ring opening with aqueous sulfuric acid, followed by acetylation with acetic anhydride (Scheme 1) [14,15]. Scheme 1. Epoxidation and hydrolysis of cycloocta-1,5-diene leading to racemic diols 2 and acetylation affording diacetates 4.
The preparation of optically active 5-cyclooctene-1,2-diol 2 was first conceived by using microwave-assisted lipase-catalyzed desymmetrization of meso-symmetric diol 2 using vinyl acetate as the acylating agent in THF as solvent rather than isooctane or 2-methylbutan-2-ol, that gave lower results. In order to perform the reaction under microwave irradiation, we decided to choose thermostable immobilized lipases capable of withstanding microwave irradiation: Novozyme 435 ® (CaLB immobilized on acrylic resin) and PS-D (Pseudomonas cepacia immobilized on diatomite). The goal was to obtain an enantioselective enhancement with immobilized lipases, as previous studies performed in our laboratory showed that resolution of rac-diol 2 with free Pseudomonas cepacia lipase at 55 °C in THF during 7 days afforded with a good conversion rac-monoacetate 3 (47%, 0% ee) and rac-diol 2 (51%, 0% ee) but with no selectivity at all. Indeed, by immobilization of enzymes onto solid supports, enhanced enzyme activity, selectivity, stability, and reusability in organic media may be achieved compared to the native enzyme [16].

Scheme 2.
Enantioselective acetylation of diol (2) using immobilized CaLB lipase and vinyl acetate by classical heating at various temperatures. Under microwave irradiation at 35 °C (5 W), racemic diol 2 proceeded to give (1R,2R)-monoacetate 3a (32%, 45% ee), trace amounts of (1S,2S)-diacetate 4b (5%, >99% ee) and 65% of diol 2 (23% ee). In order to study the influence of the irradiation power on the biocatalytic media, we decided to apply a constant power (up to 300 W) while maintaining the temperature at 35 °C by using a microwave oven combined with a Coolmate ® . We noticed an enhancement of the yield and the enantiomeric ratio of (1R,2R)-monoacetate 3a (42%, 67% ee) and diol 2 (51%, 50% ee) and only 2% of diacetate 4b (99% ee). At 50 °C (10W, 14 h) the same reaction yielded only esters: 58% of (1R,2R)-monoacetate 3a with 55% ee and 37% of diacetate 4b (99% ee). A higher temperature (80 °C, 40 W, 14 h) afforded a decrease of diacetate yield with a lower enantiomeric excess (30% of 4b with 94% ee) (Scheme 3 and Table 2).  These results suggest that, at higher temperature (100 °C), there is a loss of enzyme activity due to its denaturation. At 80 °C, Poojari et al., have shown that the immobilized-CalB (Novozym 435) was very stable and kept 90% of its activity after an incubation in diphenylether for 24 h [17]. The irradiation power appears to display a key role in enzyme properties, and best enhancement of monoacetate yield and ee was observed at 35 °C with 300 W. Brimble et al., have shown that compared to conventional heating the microwave irradiation led to higher conversion and enantiomeric excesses, in the case of lipase-catalyzed kinetic resolution of racemic secondary alcohols through acetylation [18].
The microwave-assisted hydrolysis seems to enhance the enzyme activity compared to classical heating. Comparing the results with Suemune et al. [19], the CaLB seems to be a convenient enzyme in microwave irradiation resulting in the obtention of enantiopure monoacetate 3a in a quick and clean way. However, the same reaction with PS-D needed to be optimized to obtain a better ee.

General Information
Lipases from Pseudomonas cepacia (immobilized on diatoms MKBB3465, 500 PLU −1 ) and Candida antarctica (immobilized on acrylic resin 077K1155, 10,000 PLU −1 Novozym 435 ® or free form) were purchased from Sigma Aldrich (Sigma-Aldrich Chemie S.a.r.l., Saint Quentin Fallavier, France). All other chemicals were purchased from Sigma Aldrich and were used without further purification except in the case of vinyl acetate which was used after fresh distillation.
Enantiomeric ratios were determined by gas chromatography (Agilent 7890A, Agilent Technologies SAS, Les Ulis, France) equipped with an autosampler (7688B) and flame ionization detector (FID). For the experiment, a CP-Chirasil-Dex (0.25 mm × 25 m × 0.25 µm, Chromopack, Agilent Technologies SAS, Les Ulis, France) column was used. The injector and the detector were kept at 180 °C. Nitrogen was used as gas carrier at a flow of 1.5 mL/min. Hydrogen, air and nitrogen were supplied to the FID at 35 mL·min −1 , 350 mL·min −1 and 25 mL·min −1 respectively. The products are analyzed at 110 °C. The enantiomeric ratio and yields were calculated by taking the average of two duplicates, with an error <2%.
Microwave reactions were conducted using a CEM Discover ® , single mode operating system (CEM France, Saclay, France) working at 2.45 GHz, with a programmable power ranging from 1 to 300 W. The microwave can be equipped with a Coolmate ® system allowing reactions with high power input (up to 300 W) while maintaining the reaction media at 35 °C. This system is cooled by a cryogenic fluid (Galden HT-55 ® , BT Electronics, Les Ulis, France). In closed vessel mode, microwave irradiation experiments were carried out using a single-mode microwave instrument (Initiator, Biotage, Uppsala, Sweden) working at 2.45 GHz, with a power programmable from 1 to 450 W (0-20 bars). (1) Under inert atmosphere, to a solution of cycloocta-1,5-diene (20 g, 0.161 mol) and sodium carbonate (117.6 g, 1.11 mol) in dichloromethane (560 mL) stirred at 0 °C was added dropwise a solution of peracetic acid (42.7 mL, 0.222 mol) in dichloromethane (500 mL) during 2 h. The mixture was stirred for 8 h at 0 °C and 12 h at room temperature. The mixture was quenched with 500 mL of water and extracted with (3 × 250 mL) of dichloromethane. The organic layers were dried with magnesium sulfate anhydrous and concentrated under reduced pressure. The crude residue was purified by column chromatography (silica gel, petroleum ether/ethyl acetate 90/10) to provide the (Z)-(1S,8R)-9-oxa-bicyclo[6.

Conclusions
The kinetic resolution of homochiral (Z)-cyclooct-5-ene-1,2-diols and (Z)-2-acetoxycyclooct-4-enyl acetates has been accomplished controlled using two different lipases (CaLB or PS-D) in order to obtain one or the other useful enantiomer. The role of the microwave power has also been highlighted. Finally, by microwave irradiation, this eco-efficient optimization for the resolution of racemic diols, leads to a reduction of the reaction time and a decrease of power consumption, without any toxicity.