Examples of Enzymatic Conversions

In this revised section, some recent examples have been selected emphasizing applications of enzymes to generation of natural products and derivatives and precursors combined with some of the latest technical developments. 

Due to their ready availability and the ease with which they can be handled, hydrolytic enzymes have been widely applied in organic synthesis. They do not require coenzymes, are reasonably stable, and often tolerate organic solvents. Their potential for regioselective and especially for enantioselective synthesis makes them valuable tools [10, 14, 16, 19]. 

Regioselective acylations of polyhydroxylated compounds such as carbohydrates, glycerols, steroids, or alkaloids have been carried out with lipases, esterases, and proteases [13, 20]. One example is the Candida antartica lipase (immobilized on acrylic resin) catalyzed monoacylation of the signalling steroid ectysone (1) giving selectively the 2-C)-acetate 2 (eq. (1)). Using vinyl acetate for this transesterification the reaction was irreversibly pushed to the product side, since the liberated enol instantaneously isomerizes to acetaldehyde [21]. The sometimes unfavorable aldehyde is avoided when 1-ethoxyvinyl acetates [22], trichloro- or -fluoroethyl esters [23 a, b], oxime esters [23 c] or thioesters [23 d] are employed for the quasi-irreversible reaction courses. 

In the synthesis of highly phosphorylated phosphoinositide derivatives regioselective hydrolysis of one out of three butyrates by a lipase in pH 7.8 buffer containing 

Pig liver esterase (PLE) has found extensive use for the hydrolytic cleavage of methyl or ethyl esters of prochiral carboxylic acids. 

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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... 

As most enzymes function under compatible ambient conditions, bio-bio cascades had already been successfully developed by the 1970s. By far, most examples have been reported in the field of carbohydrates, using combinations of enzymatic conversions (up to eight enzymes in one-pot), as well as for the in situ cofactor regeneration of enzymatic redox reactions towards amino and hydroxy acids. 

The interest in catechol oxidase, as well as in other copper proteins with the type 3 active site, is to a large extent due to their ability to process dioxygen from air at ambient conditions. While hemocyanin is an oxygen carrier in the hemolymph of some arthropods and mollusks, catechol oxidase and tyrosinase utilize it to perform the selective oxidation of organic substrates, for example, phenols and catechols. Therefore, establishment of structure-activity relationships for these enzymes and a complete elucidation of the mechanisms of enzymatic conversions through the development of synthetic models are expected to contribute greatly to the design of oxidation catalysts for potential industrial applications. 

Here we confine ourselves to a few typical examples of enzymatic systems in organic solvents (Klibanov, 2001 and references therein), a-chymotripsin is stable in unhydrous conditions for several hours at 100°C. The a-chymotrypsin hydrophilic peptide substrate is transformed in organic solvent three times faster than hydrophobic substrate, while the latter in water is found to be non-reactive In water solution the dominant product of the conversion ofprochyral 2-(3,5-dimetoxybenzyl)l,3-propandiol by this enzyme is the S-monoester, whereas in acetonitril R-enantiomer is formed. 

A biocatalytic system for converting biomass to industrial chemicals is not only applicable to enzymatic conversions but also to fermentative conversion using cellulose. We report here three examples of fermentative conversion of cellulose to chemicals namely 1,3 propanediol, lactic acid, and succinic acid. 

Two examples of enzymatic resolutions with selectivities of = 5 and = 20 are depicted in Fig. 2.4. The curves show that the product (P -E Q) can be obtained in its highest optical purities before 50% conversion, where the enzyme can freely choose the weU-fitting enantiomer from the racemic mixture. So, the well-fitting enantiomer is predominantly depleted from the reaction mixture during the course of the reaction, leaving behind the poor-fitting counterpart. Beyond 50% conversion, the enhanced relative concentration of the poor-fitting counterpart leads to... 

The MER concept has successfully been applied to a series of enzymatic conversions, each of which is useful in analysis, diagnostics, high-throughput screening, or reaction analysis as pointed out in the following examples. 

Fructose—Dextrose Separation. Emctose—dextrose separation is an example of the appHcation of adsorption to nonhydrocarbon systems. An aqueous solution of the isomeric monosaccharide sugars, C H 2Dg, fmctose and dextrose (glucose), accompanied by minor quantities of polysaccharides, is produced commercially under the designation of "high" fmctose com symp by the enzymatic conversion of cornstarch. Because fmctose has about double the sweetness index of dextrose, the separation of fmctose from this mixture and the recycling of dextrose for further enzymatic conversion to fmctose is of commercial interest (see Sugar Sweeteners). 

Increasingly, biochemical transformations are used to modify renewable resources into useful materials (see Microbial transformations). Fermentation (qv) to ethanol is the oldest of such conversions. Another example is the ceU-free enzyme catalyzed isomerization of glucose to fmctose for use as sweeteners (qv). The enzymatic hydrolysis of cellulose is a biochemical competitor for the acid catalyzed reaction. 

Many products made by fermentation are also based on the conversion of starch. Some examples of the use of enzymatically hydrolyzed starches are the production of alcohol, ascorbic acid, enzymes, lysine, and penicillin. 

A number of examples of monoacylated diols produced by enzymatic hydrolysis of prochiral carboxylates are presented in Table 3. PLE-catalyzed conversions of acycHc diesters strongly depend on the stmcture of the substituent and are usually poor for alkyl derivatives. Lipases are much less sensitive to the stmcture of the side chain the yields and selectivity of the hydrolysis of both alkyl (26) and aryl (24) derivatives are similar. The enzyme selectivity depends not only on the stmcture of the alcohol, but also on the nature of the acyl moiety (48). 

Almost all types of cell can be used to convert an added compound into another compound, involving many forms of enzymatic reaction including dehydration, oxidation, hydroxyla-tion, animation, isomerisation, etc. These types of conversion have advantages over chemical processes in that the reaction can be very specific, and produced at moderate temperatures. Examples of transformations using enzymes include the production of steroids, conversion of antibiotics and prostaglandins. Industrial transformation requires the production of large quantities of enzyme, but the half-life of enzymes can be improved by immobilisation and extraction simplified by the use of whole cells. 

The use of high performance liquid chromatography (HPLC) for the study of paralytic shellfish poisoning (PSP) has facilitated a greater understanding of the biochemistry and chemistry of the toxins involved. HPLC enables the determination of the type and quantity of the PSP toxins present in biological samples. An overview of the HPLC method is presented that outlines the conditions for both separation and detection of the PSP toxins. Examples of the use of the HPLC method in toxin research are reviewed, including its use in the determination of the enzymatic conversion of the toxins and studies on the movement of the toxins up the marine food chain. 

Cascade catalysis without recovery of intermediate products may require more than two steps, involving enzymatic, homogeneous, and heterogeneous catalysis. Several examples of this approach have been given [72, 73] one of the most representative consists of a four-step conversion of glucoside into aminodeoxysugar without intermediate product recovery. 

After some early examples of bio-chemo combinations in the 1980s, there was then over a decade of silence , followed by clearly increasing interest from the mid-1990s in the field of dynamic kinetic resolution processes (i.e., chemocata-lyzed racemization combined with enantioselective enzymatic conversion, giving, in principle, 100% yield of an optically pure compound). 

Apart from the impressive recent examples given above, however, there has been too little focus thus far on developing a toolkit of chemocatalytic conversions that are as mutually compatible as enzymatic reactions are in nature (presently there are great differences in solvent, temperature, sensitivity to air and moisture, reactants). This confirms in fact the main difference in approach between organic synthesis and biosynthesis organic synthesis employs a maximum diversity in reagents and conditions while biosynthesis exploits subtlety and selectivity from a small range of materials and conditions (Fig. 13.7). 

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