Enzymatic routes

Padhi, S.K., Kaluzna, I.A., Buisson, D. et al. (2007) Reductions of cyclic beta-keto esters by individual Saccharomyces cerevisiae dehydrogenases and a chemo-enzymatic route to (lR,2S)-2-methyl-l-cyclohexanol. Tetrahedron Asymmetry, 18 (18), 2133-2138. 

Groger, H. (2001) Enzymatic routes to enantiomerically pure aromatic a-hydroxy carboxylic acids a further example for the diversity of biocatalysis. Advanced Synthesis and Catalysis, 343, 547-558. 

Figure 14.1 Scheme for (a) chemical and (b) enzymatic routes for production of cephalexin... 

In vitro polyester syntheses using an isolated enzyme as catalyst via non-bio-synthetic pathways is reviewed. These enzymatic routes for production of biodegradable polyesters possess several advances in comparison with fermentation and chemical processes ... 

Several enzymatic routes with sucrase-type enzymes have been established for oligosaccharide synthesis, some of which are applied industrially for food and feed ingredients, and important examples are compiled.5 Convenient routes for new oligosaccharides, however, are rarely available. 

As would be expected, the chemistry is complex. Unpleasant, off flavor odors usually derive from sulfur compounds, such as MT, DMS, DMDS, and DMTS, formed either enzymatically or non-enzymatically from sulfur-containing amino acids.35 Enzymatic routes to MT are essentially those previously considered (Section 11.1.2.4.5). Some DMS may derive by methylation of MT (Equation 8) with the donor, 5-adenosylmethionine, AdoMet ... 

Organic phosphates, phosphonates, and analogues are of interest in medicinal chemistry as drugs or prodrugs, and in toxicology mainly as plasticizers, insecticides, or warfare agents. Their breakdown by enzymatic routes has received marked attention since many years, but it is most unfortunate for the advancement of science that an unknown number of highly relevant results remain classified information. 

Dunsmore, C.J., Carr, R., Fleming, T. and Turner, N.J., A chemo-enzymatic route to enantio-merically pure cyclic tertiary amines. J. Am. Chem. Soc., 2006, 128, 2224. 

Enzymatic reduction of carbonyl compounds and enzymatic enantioselective transformation of racemic or meso alcohols (25,43.) are two methodologies that have proven to be beneficial in the preparation of optically active hydroxyl compounds, key chiral building blocks used in carbohydrate and natural product syntheses (44-45. Our interest in this area is to develop enzymatic routes to optically active glycerol and furan derivatives, and hydroxyaldehydes. 

The energetics of enzymatic and their corresponding uncatalyzed reference reactions can be understood by the cyclic path that allows for substrate conversion to product by the uncatalyzed and enzymatic routes (Fig. 2). Note that the uncatalyzed reaction is characterized by a transition state that is far less stable than its enzymatic counterpart. Note also that the initial and final conditions are the same for either route, an absolute requirement for any catalyzed process i.e., no effect on the overall equilibrium constant). 

Here, we will focus on the enzymatic routes since enzymatic preparation of DHAP is usually coupled with the aldol addition catalyzed by the aldolase representing genuine multi-enzyme systems. 

Scheme 4.5 Enzymatic routes to DHAP based in the use of glycerophosphate oxidase (GPO) coupled with the aldolase-catalyzed reaction and with dephosphorylation of the aldol adduct.

The scheme shown in Figure 1 is complicated, and simpler explanations to account for the difficulties encountered to date in demonstrating cytosolic isozymes are a logical first course of action. Possibilities currently under evaluation are (i) Most cytosolic isozymes may be particulate (ii) Different enzymatic routes may be employed in the cytosol (iii) Detection of cytosolic isozymes may require conditions of secondary metabolism (e.g., after wounding) that maximize expression levels of enzyme and (iv) Detection of cytosolic isozymes may require conditions of normal growth physiology that maximize expression levels of enzyme. 

Figure 2. Alternative enzymatic routing for L-phenylalanine biosynthesis. Dehydration followed by transamination defines the phenylpyruvate route, whereas the reverse order of reactions defines the arogenate route. Abbreviations GLU, L-glutamate aKG, 2-ketoglutarate.

Verschraagen, M., Boven, E., Torun, E., Hausheer, F. H., Bast, A., and van der Vijgh, W. J. F., Possible (enzymatic) routes and biological sites for metabohc reduction of BNP7787, a new protector against cisplatin-induced side-effects. Biochemical Pharmacology 68(3), 493-502, 2004. 

Fig. 1 Enzymatic routes (a-c) of telechelic diacrylates by enzymatic ROP for subsequent crosslinking. X denotes the polymerizable (meth)acrylate group...

Scheme 6 Enzymatic routes (a and b) towards graft copolymers...

Scheme 2.2.5.S Multi-enzymatic route for the stereoselective synthesis of D-xylulose 5-phosphate 19 from commercial D-fructose 1,6-bisphosphate (abbreviations TPI, triose phosphate isomerase TK, transketolase).

The enzymatic route which a drug or poison follows in its metabolism is very specific to the xenobiotic itself. Substances with the same type of structures need not go through the same pathway. And the "active" form of the drug or the "toxic" form of the poison may occur either at the beginning or during the course of transformation. Usually synthetic combinations are not active or toxic but many of their precursors are. The following diagram illustrates the possibilities of a biotransformation process. 

The examination of the total reagent usage in the two processes (last row in Table 8.3) clearly shows that the new enzymatic route (with recycling of 4) utilizes 7 times less input of chemicals. This includes 12 times less input of solvents as compared to the first-generation route. Moreover, in the optimized process, every chemical reaction is run in water with minimal solvents used for work-up. Some of the process water can be sent directly to the wastewater treatment plant, and the solvent from the hydrolysis/decarboxylation process is recovered. Further improvements from pilot plant and production scale runs have been demonstrated and will be implemented in the future. 

The overall conversion of 4 to API was thus progressively improved from 25.8% for the classical route to 33.4% for the enzymatic route (no recychng) and 42% for the enzymatic route with recychng. Pregabahn is a large-volume product for Pfizer. Based upon projected sales and the figures in Table 8.3 (final column), it is expected that between 2007 and 2020 the environmental savings (versus those in the classical-resolution column) wiU be ... 

Table 8.3 Key material inputs for classical resolution and enzymatic routes.

Classical resolution route (Scheme 8.2) Enzymatic route (Scheme 8.6) ... 

Table 8.4 Energy usage results for first-generation and enzymatic routes. ...

Stage Classical resolution route Energy (MJ/kg 1) Enzymatic route (no recycle of 4) Energy (MJ/kg 1)... 

The enzymatic route resulted in an energy usage reduction of 82% versus the classical resolution process. This energy usage was based on two main factors (i) 2) Batch Plus version 2006.5 by Aspen Technology. 

Several chemical and enzymatic methods are available for the synthesis of glycosyl phosphates. The required nucleoside triphosphates (NTPs) are most conveniently prepared by enzymatic routes. In general, these methods involve the sequential use of two kinases to transform NMPs to NTPs, by the corresponding NDPs (Fig. 2, see a recent review [12] for more details). 

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