Читать книгу Enzyme-Based Organic Synthesis - Cheanyeh Cheng - Страница 25

Since primary alcohols can be oxidized to aldehydes and further to carboxylic acids, which are versatile building blocks in organic synthesis, the selective oxidation of primary alcohols using enzymes to produce corresponding aldehydes is important for both fundamental and industrial research. Benzaldehyde, which is important and widely applied in cosmetic, flavor, and pharmaceutical industries [2, 3], is generally prepared by oxidation of toluene or hydrolysis of benzyl chloride. Due to environmental concern, scientific reports have shown that Gluconobacter oxydans can be used for the selective oxidation of benzyl alcohol to obtain benzaldehyde in an organic/aqueous biphasic system to substitute for these environmentally unfavorable processes [4, 5]. Optimization of the immobilization parameters for G. oxydans not only improves the bioconversion of the selective oxidative process but also increases the stability of immobilized cells so that the immobilized cells can be used repeatedly for 10 cycles and a 53.2% of the oxidative activity of the immobilized cells compared with that of free cells was retained [6].

Fourteen commercial alcohol oxidases, 33 selected commercial alcohol dehydrogenases (ADHs), and 218 microorganisms were tested for the oxidation of benzyl alcohol for the production of benzaldehyde via different types of hydrogen transfer [7]. If alcohol oxidases were used for the oxidation, the reaction just required molecular oxygen as oxidant and hydrogen peroxide was produced as a side product, which subsequently is decomposed disproportionately by a catalase to form water and molecular oxygen (Scheme 2.1). If isolated ADHs or lyophilized microorganisms were employed for the oxidation of benzyl alcohol, the hydrogen acceptor of the hydrogen transfer reaction could be acetaldehyde (Scheme 2.2), chloroacetone, or acetone.

Direct oxidation of simple primary alcohols such as methanol and ethanol to corresponding aldehydes using either free whole cell or immobilized whole cell was also reported in literature. For methanol oxidation to formaldehyde, methanol oxidase (MOX) in the yeast Hansenula polymorpha was involved for the biotransformation [8]; for ethanol oxidation to acetaldehyde, ADH in Saccharomyces cerevisiae was employed for the biotransformation. In these studies, the immobilization of microbial cells was found to be of great help for both the cell activity and stability [9].

Scheme 2.1 Oxidation of benzyl alcohol using an oxidase and a catalase.

Scheme 2.2 Oxidation of benzyl alcohol via alcohol dehydrogenases or microbial cells.

The stereoselective oxidation of secondary alcohols to produce ketones is of greatest interest in organic synthesis for its applications in pharmaceutical industries. Simple sec‐alcohol, 2‐butanol, has been oxidized to butanone by the immobilized yeast S. cerevisiae with a 45% yield [9]. Three screened yeast, Williopsis californica, Williopsis saturnus, and Pachysolen tannophilus, have been used for the oxidation of six cycloalkanols with different ring size including cyclobutanol, cyclopentanol, cyclohexanol, cycloheptanol, cyclooctanol, and cyclododecanol. The results show that W. californica and P. tannophilus are active against all six cycloalkanols and can be thought as nonselective, while W. saturnus is active against cycloalkanols of four, five, and six carbon atoms and is selective for small cyclohexanols [10]. These three selected strains have also been employed for exploring the stereoselectivity of several sec‐alcohols such as (1R)‐(2‐furyl)‐ethanol, (1S)‐(2‐furyl)‐ethanol, (1R)‐phenyl‐ethanol, (1S)‐phenolethanol, (1R)‐tetrahydronapthol, (1S)‐tetrahydronapthol, (−)‐neo‐menthol((1R,2R,5S)‐2‐isopropyl‐5‐methyl‐cyclohexanol),(+)‐menthol ((1S,2R,5S)‐2‐isopropyl‐5‐methyl‐cyclohexanol), and iso‐menthol ((1S,2R,5R)‐2‐isopropyl‐5‐methyl‐cyclohexanol). The results indicate that all the strains are stereoselective toward the S‐enantiomer [10].

The application of primary alcohol oxidation is proved to be valuable for the preparation of enantiopure 1,2‐diols [11]. Several enantiopure 1,2‐diols were produced via a simple and green regio‐ and stereoselective concurrent oxidation of the racemates using Sphingomonas sp. HXN‐200 (Scheme 2.3). For the four tested 1,2‐diols, this concurrent oxidation process demonstrates better enantioselectivity than any other known biocatalytic processes.

Another method used for the preparation of enantiopure (S)‐1‐phenyl‐1,2‐ethanediol from the racemic (R,S)‐1‐phenyl‐1,2‐ethanediol was performed with the whole‐cell yeast Candida parapsilosis. In this method, (R)‐1‐phenyl‐1,2‐ethanediol is first stereoselectively oxidized to 2‐hydroxyacetophenone by a (R)‐specific ADH, and subsequently the intermediate is stereoselectively reduced by an (S)‐specific carbonyl reductase to form (S)‐1‐phenyl‐1,2‐ethanediol (Scheme 2.4). The catalytic activities of the two key enzymes in C. parapsilosis responsible for the deracemization were significantly enhanced by the increase of dissolved oxygen concentration with higher agitation speed [12].

Scheme 2.3 Regio‐ and stereoselective concurrent oxidations of (±)‐1,2 diols.

Scheme 2.4 Deracemization of racemic 1‐phenyl‐1,2‐ethanediol by C. parapsilosis through an oxidation–reduction sequence.

Enantiopure sec‐alcohols and α‐hydroxyketones (acyloins) are popular and important synthons for the asymmetric synthesis of many bioactive compounds [13–15]. A biocatalytic racemization is proved to be a useful tool for the preparation of enantiopure simple alcohols and acyloins via an enzymatic oxidation–reduction process with the formation of a nonchiral ketone or α‐diketone intermediate, respectively, using whole lyophilized cells of various bacteria, fungi, and one yeast [16]. The kinetic resolution process was applied for three selected sec‐alcohols (2‐octanol, 6‐methyl‐5‐heptene‐2‐ol, and 1‐phenylethanol) and three selected acyloins (benzoin, phenyl acetylcarbinol, and 2‐hydroxy‐1‐indanone) in which phenyl acetylcarbinol and derivatives are key intermediates for industrially manufacturing (−)‐(pseudo)ephedrine, antidepressants, and smoking cessation agents [17–20], and 2‐hydroxy‐1‐indanone is used for the synthesis of several human immunodeficiency virus (HIV)‐protease inhibitors [21, 22]. Sec‐alcohol dehydrogenases and α‐diketone reductases are responsible for the equilibrium‐controlled enzymatic oxidation–reduction sequence, respectively, and the racemization protocol gives (S)‐enantiomers (Scheme 2.5).

Glucose dehydrogenases, glucose oxidases, and gluconate dehydrogenases have been exploited for the oxidation of aldoses on large‐scale applications and biosensors [1]. For instance, glucose oxidase (GO_x) has been widely employed in enzymatic kits for fast glucose determination and for the industrial production of gluconic acid from glucose, which was carried out using whole cells of Aspergillus niger. The industrial production of gluconic acid from glucose can be also performed by a membrane‐bound dehydrogenase from G. oxydans. A more recent industrial application of GO_x is the direct oxidation of D‐glucosamine to D‐glucosaminic acid with the combined use of catalase to degrade the H₂O₂ produce.

Scheme 2.5 Biocatalytic racemization of sec‐alcohols and acyloins using lyophilized microbial cells.

Direct oxidation of heterocyclic and aromatic aldehydes to the corresponding carboxylic acids can be accomplished by Acetobacter rancens IFO3297, Acetobacter pasteurianus IFO13753, and Serratia liquefaciens LF14 [1, 23]. For instance, oxidation of furfural by A. rancens IFO3297 can produce 110 g L⁻¹ of 2‐furoic acid with a 95% yield. 5‐Hydroxymethyl‐2‐furancarboxylic acid obtained from corresponding aldehyde can be obtained by whole cells LF14. Isophthalaldehyde, 2,5‐furandicarbaldehyde, 2,5‐thiophenedicarbaldehyde, and 2,2′‐biphenyldicarbaldehyde can be converted to the corresponding formylcarboxylic acid with 86–91% yields by both IFO13753 and LF14. The aromatic carboxylic acids such as vanillic acid, p‐hydroxybenzoic acid, and syringic acid can be produced by the oxidation of corresponding aromatic aldehydes using whole‐cell Burkholderia cepacia TM1 [1, 24].

The oxidation of alcohols and aldehydes to form corresponding aldehydes, ketones, and carboxylic acids has also been used by cells to produce important intermediates or precursors during the biosynthetic pathways. In the biosynthesis of depside atranorin from acetate using immobilized lichen cells Evernia prunastri, the most probable precursor, an aldehyde‐substituted phenolic aldehyde, haematommic acid, is produced by oxidation of an alcohol intermediate as shown in Scheme 2.6 [25]. In another example, resting cells of Nocardia iowensis DSM 45197 have been selected for the synthesis of vanillin and vanillic acid from the starting material isoeugenol. Three possible pathways used by N. iowensis have been suggested for the conversion of isoeugenol to vanillic acid and vanillin. ¹⁸O‐labeling studies showed that most likely route appears to be the initial side‐chain olefin epoxidation of isoeugenol, epoxide hydrolysis to a vicinal diol, followed by diol cleavage to vanillin and subsequently oxidation of the aldehyde group of vanillin to vanillic acid by an aldehyde oxidase (Scheme 2.7) [26].

Scheme 2.6 Oxidation of an alcohol intermediate to the precursor of atranorin in the biosynthetic pathway of lichen cells.

Scheme 2.7 Bioconversion of isoeugenol to vanillin and vanillic acid by N. iowensis.