Industrial biotransformations isolated enzymes

Some of the industrial biocatalysts are nitrile hydralase (Nitto Chemicals), which has a productivity of 50 g acrylamide per litre per hour penicillin G amidase (Smith Kline Beechem and others), which has a productivity of 1 - 2 tonnes 6-APA per kg of the immobilized enzyme glucose isomerase (Novo Nordisk, etc.), which has a productivity of 20 tonnes of high fmctose syrup per kg of immobilized enzyme (Cheetham, 1998). Wandrey et al. (2000) have given an account of industrial biocatalysis past, present, and future. It appears that more than 100 different biotransformations are carried out in industry. In the case of isolated enzymes the cost of enzyme is expected to drop due to an efficient production with genetically engineered microorganisms or higher cells. Rozzell (1999) has discussed myths and realities... 

The use of isolated purified enzymes obviously avoids the formation of unwanted byproducts which may be generated from otherwise present and usually unknown enzymes. Isolated enzymes may be more active and selective than whole cells, but their recovery and purification is very expensive. Whole cell biotransformations show a lot of advantages such as the increased stability of enzymes which exist in whole cells. If these enzymes depend on cofactors, whole cell biotransformations become even more favorable because the use of whole cells allows the intracellular cofactor pool to be exploited, and any addition of exogenous cofactors becomes unnecessary. Otherwise the costs for the required cofactors often exceed that for the enzyme or the value of the product. Therefore, whole cell biotransformations are very interesting for industrial applications. 

The application of isolated enzymes to preparative organic synthesis on an industrial scale is a matter of active research worldwide. Since the late sixties, immobilized enzymes have been used in amino acid production in continuous processes on a large scaled 2. In the late seventies, the use of soluble enzymes, especially in membrane reactors, broadened the scope of enzyme technology13, 41 and opened the way to simultaneous use of more than one enzyme for complex conversions -especially coenzyme-dependent biotransformations[5-7. In the early 1980s the use of enzymes was extended further to involve organic solvents[8, 9 ... 

Biotransformations involve the use of isolated enzymes or intact microbial cells for the highly selective transformations of organic molecules for cutting edge preparative organic syntheses. Ideally, the use of biocatalysts (i.e. enzymes or whole cells) in the industrial preparation of useful compounds would be economical and environmentally friendly. Readers wishing a more thorough discussion on this topic are referred to a recently published article [85]. 

Biotransfonnations are selective chemical modifications of molecules by biological entities ranging from isolated enzymes (ceU-free systems) to intact organisms. In particular cases they are exploited by the chemical and pharmaceutical industries for production of chemicals this approach can be especially advantageous for structurally complex compounds. Preparative biotransformations have been used on a laboratory scale by radiochemists to prepare isotopically labeled compounds necessary for drug development and other applications in the life sciences. Their synthetic use for isotope labeling includes ... 

Though use of isolated purified enzymes is advantageous in that undesirable byproduct formation mediated by contaminating enzymes is avoided [37], in many industrial biotransformation processes for greater cost effectiveness the biocatalyst used is in the form of whole cells. For this reason baker s yeast, which is readily available, has attracted substantial attention from organic chemists as a catalyst for biotransformation processes. One of the first commercialized microbial biotransformation processes was baker s yeast-mediated production of (R)-phenylacetyl carbinol, where yeast pyruvate decarboxylase catalyzes acyloin formation during metabolism of sugars or pyruvate in the presence of benzaldehyde [38]. 

This chapter gives a brief review of the isolation and production of enzymes. More detailed information can be obtained from various published textbooks and reviews11"51. Most of the industrial enzymes used for chemical synthesis are supplied in a crude form with an active enzyme content of only a few percent. The other constituents are inorganic salts, polysaccharides and diatomaceous earth used as stabilizers and excipients. Purified enzymes for biotransformation are supplied by some manufacturers in a crystal or immobilized form. These enzymes, though expensive, are easy to apply for biotransformation in organic media. The use of more purified enzymes is increasing. 

The quest for microorganisms capable of performing the desired biotransformation of indene led to the isolation of several strains of the genus Rhodococcus from soil samples contaminated with aromatic compounds that are able to oxidize indene to 1,2-indandiols of different chirality, and various other oxygenated derivatives [8]. Induction studies indicated that several oxygenases were present and differentially induced by naphthalene, toluene, and indene. The stereospecific nature of the enzymes expressed in Rhodococcus as well as their abihty to tolerate indene as a substrate makes these microorganisms promising candidates for development as an industrial-scale biocatalyst for the production of (21 )-indandiol. 

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