Organic enzymatic methods

Aldoximes are prepared from aldehydes and hydroxylamine by condensation reaction, and the dehydration reaction of aldoxime is one of the most important methods of nitrile synthesis in organic chemistry." We speculated that it would become one of the most important examples in Green Chemistry if the dehydration reaction could be realized by an enzymatic method, and started studies on a new enzyme, aldoxime dehydratase, and its use in enzymatic nitrile synthesis. Furthermore, we clarified the relationship between aldoxime dehydratase and nitrile-degrading enzymes in the genome of the microorganisms and the physiological role of the enzyme. 

Because of the need for isolation techniques applicable to other species, Hoogenboom et al [18] investigated the isolation of hepatocytes from porcine livers and developed an enzymatic method with closed-loop and open-loop perfusion stages for the perfusion of liver lobes. We investigated a method to perfuse the whole organ to receive a great amount of hver cells (up to 25-10 cells per pig liver) and a viability higher than 90%. 

Another interesting example of resolution through formation of diastereo-mers is the isolation of four stereoisomers of 3-amino-2-methyl-3-trifluoro-methyl butanoic acid [55]. In this process, the chemical-enzymatic method by the combination of chemical and enzymatic reaction is a very convenient. At first, -phenylacetyl derivatives 61a and 61b were prepared in excellent isolated yields via the Schotten-Baumann procedure. After these materials were hydrolysed with penicillin acylase (EC 3.5.1.11) from Escherichia coli until attainment of 50% conversion, enzymatically unconverted -phenylacetyl derivatives 62 a and 62 b (organic layer) and amino acids 63 b and 63 d (aqueous layer) were separated. Acidic hydrolysis of unconverted materials produced other stereoisomers 63 a and 63 c in high optical pure form. 

Many methods have been used to determine organic acids in foods, including volumetric, electrochemical, enzymatic, and chromatographic (paper, thin-layer, gas-liquid, or HPLC) methods. However, most of these are not able to assay organic acids comprehensively for example, the enzymatic methods are specific kits for individual organic acids (i.e., they only detect one of the acids present). Therefore, to analyze the sample comprehensively using enzymatic methods would be extremely time consuming and costly, as the analysis would have to be run separately for each acid and would require several kits. 

ISO (2006b). Cheese and Processed Cheese Products—Determination of Citric Acid Content—Enzymatic Method (Standard ISO/TS 2963/IDF/RM 34). International Organization for Standardization, Geneva. 

Methods for chemical analysis of NAD(P)H include enzymatic methods [37] and bioluminiscence methods ([38] by Grupe and Gottschalk using a commercial test kit from Boehringer Mannheim). Increased NADH levels often indicate oxygen limited states of aerobic organisms (well known for baker s yeast, and also observed with hybridoma cultures by Zupke et al. [39], where the situation is more complex due to cell compartments). 

Enantioselective Intramolecular Cyclization (Sn2 reaction). The desymmetric transformation of meso-structures has been recognized as a versatile synthetic method for optically active compounds in organic enzymatic processes. The enantioselective intramolecular cyclization of the bis-phenyllithium species, which is generated by addition of butyllithium to a solution of cis-3,5-di(bromophenoxy)cyclopentene, has been attained by addition of lithium salt (1.2 equiv) of (/ )-BINOL-Me to produce a cyclopenta[fc]benzofuran with 87% ee (eq 18). ... 

Asymmetric synthesis is a very important area in organic chemistry. However, it must be noted that electrochemical asymmetric synthesis has been immature compared with chemical, catalytic, and enzymatic methods, although much effort has been made since the first report in this area by Grimshaw and coworkers [414] in 1967. 

The enantioselective discrimination of one of the hydroxyl groups of meso-diols can give chiral monoprotected diols, which serve as versatile intermediates for asymmetric organic synthesis. In addition to the enzymatic methods, a number of chemical approaches have been reported using chiral 1,2-diamine catalysts, chiral phospholane-based catalysts, planar chiral DMAP derivatives, and oligopeptide-based catalysts [2,28], Surprisingly, however, relatively a few publications are devoted to this reaction with cinchona-based organocatalysts. 

Several other organic acids, especially lactic, are present in cheese, and are routinely analyzed by HPLC. Enzymatic methods are available for several acids, e.g., D and L lactic (the easiest method available to distinguish between the isomers), acetic, pyruvic, and succinic (Boehringer-Maimheim, 1986). 

Several enzymatic methods in conjunction with conventional organic methods provide new synthetic routes to fluorinated sugars. These new methods allow preparative synthesis of sugars containing the fluorine probe for use in the NMR study of receptor-substrate interactions of enzymes, of cell-surface glycoproteins, and of other systems related to biochemical recognition... 

Anthracyclines are large and chemically labile molecules, which makes their synthetic modification challenging, hi biosynthetic methods, reactions are performed mostly in neutral aqueous solutions at moderate temperatures. Enzymatic reactions are highly specific for the position as well as the stereoisomer modified Hkewise, chiral products are exclusively of one stereoisomer. Particularly in vivo, in Hving organisms, enzymatic reactions can be combined to perform several modifications in one process, or, indeed, to build the entire molecule from simple precursors. One of the strengths of biosynthetic methods is also their main weakness only modifications for which an enzyme exists can be performed. 

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