Use of Enzyme Preparations

Competition and trapping experiments make it possible to study hypothetical intermediates even if they are not available in an isotopically labeled form. The experiments fail, however, if endogenous intermediates are strictly channeled and do not mix with compounds administered from outside (A 3). 

Methodology, Specialist Periodical Report. Biosynthesis i, 1-40 (1972) 

Buhner, M., Schmidt, L. Die Synthese Kohlenstoff-14-markierter organischer Verbindungen, Thieme, Leipzig 1966 

Muramatsu, M. (eds.) Radiotracer Techniques and Application. Decker, New York-Basel 1977 

Instrumental methods in biosynthetic studies. Lloydia 35, 399-417 (1972) Grutzner, J. B. Carbon-13 NMR spectroscopy and its application to biological systems. Lloydia 35, 375-398 (1972) 

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Dimitroff, D. and Prodanski, P. 1973. Use of enzyme preparations of microbial origin in kachkaval cheese manufacture. Production of ewes and cows milk kachkaval cheese using an enzyme preparation from Bacillus mesentericus. Milchwissen-schaft 28, 568-571. 

Significant further progress in marine biosynthesis will be achieved when methods are developed to perform stable isotope incubations with cell-free systems of marine invertebrates. The use of enzyme preparations provides a convenient and rapid method of evaluating biosynthetic precursors, and obviates problems associated with transport of prospective intermediates across cell membranes. 

This nonreducing disaccharide, a-D-glucopyranosyl a-D-glucopy-ranoside, was enzymically synthesized by Cabib and Leloir (with yeast-enzyme preparation) from UDP-D-glucose plus D-glucose 6-phosphate it was obtained as a phosphorylated derivative, namely, a,Q -trehalose 6-phosphate. In a subsequent step, the phosphate group was hydrolyzed oflF by a phosphatase, affording the free disaccharide. The formation of a,a-trehalose was similarly shown to ensue from the same substrates by use of enzymic preparations from insects. ... 

One of the major obstacles in making full use of Na /K -ATPase in primary screening is the fact that the enzyme preparations from human cardiac muscle and brain cortex consist of a mixture of isoenzymes [61], the cellular origins, the proportions, and the properties of which are still unknown. The physical separation of pure isoenzymes appears at present to be inaccessible. In the meantime, several ways out of this dilemma may be tried. These include the use of enzyme preparations from kidney or skeletal muscle, which are known to express only the cxl isoenzyme or preferentially the al isoenzyme, respectively [61]. 

The use of enzyme preparations for the elucidation of secondary metabolic pathways is described in B 3. Some general principles of reactions catalyzed by the most important groups of secondary metabolic enzymes are given in Chapter C. 

Antunes, A.F. dos S. (1978) Use of enzyme preparations in olive oil extraction. Bol Inst. Azeite Produtos Oleaginosa 6, 39-52. 

By the use of enzyme preparations from trjrptophan-adapted cells of Pseudomonas, these investigators obtained the conversion of kynurenic acid to glutamic acid. Study of the intermediate steps from kynurenic acid with separated enzyme fractions by Hayaishi and co-workers 360, 851, also private communication) led to the formulation of the scheme of kynurenic acid dissimilation shown in Fig. 21. 

A. tamurri. The qualities of the leathers manufactured by dehairing with the enzyme depilant developed from A. tamurri and by lime - sulphide method have been comparatively assessed with respect to their physical parameters. Pal et al. (1996) has recommended the use of enzyme preparations from Rhizopus oryzae for dehairing. 

The influence of enzyme maceration using pectinolytic enzyme preparations (Pectofruit and Pectofruit Press) on anthocyanin extraction at 43°C from two variants of black currant berries was studied. Enzymes accelerated the extraction yield the yield of anthocyanin extraction was similar for both enzymes and the duration did not influence the total content of released pigments. 

Until recently, the only marine example of cycloartenol (32) production was in the chrysophyte Ochromonas sp. [20], A survey, documenting the products of squalene oxide (37) cyclization (see Scheme 3) using crude enzyme preparations of various algal phyla has recently been reported [21]. Interestingly, while all... 

The cholinesterase to determine the toxic activity may be chosen (i) in pure form of commercial enzyme from animals in a water buffer solution or using biosensors, enzyme preparation impregnated into a rigid matrix that significantly activates the enzymic activity and (ii) in the form of crude extracts from plant or animal tissues. 

For a while, in the early 1990s, the interest in the use of enzymes in organic synthesis increased at an almost exponential rate and two-volume works were needed even to summarize developments in the field151. Now, at the turn of the century, it is abundantly clear that the science of biotransformations has a significant role to play in the area of preparative chemistry however, it is, by no stretch of the imagination, a panacea for the synthetic organic chemist. Nevertheless, biocatalysis is the method of choice for the preparation of some classes of optically active materials. In other cases the employment of man-made catalysts is preferred. In this review, a comparison will be made of the different methods available for the preparation of various classes of chiral compounds161. 

There are important developments to be expected in the commercial production of PM. As noted above, this enzyme proved to be a useful demethylating agent in the production of low-ester pectins. The use of enzymes has certain advantages over acid or alkali, and this fact might eventually create considerable market for PM preparations that are free or essentially free of PG. 

At first sight, it appears that it should be feasible to prepare such esters regioselectively using a similar biocatalytic approach to that employed for the 6- and 7-amino acylation of 6-APA and 7-ADCA shown above. Unfortunately, owing to the poor nucleophilicity of alcohols, biocatalytic esterification in aqueous media is far more challenging than amida-tion. Therefore, it was not until the pioneering work of Klibanov and co-workers, who first demonstrated the use of enzymes in neat organic solvents, that this option became viable (see Section 1.4). 

One of the first applications of the microbial hydrolysis of epoxides for the synthesis of a bioactive compound is based on the resolution of a 2,3-disub-stituted oxirane having a cis-configuration (Scheme 14). Thus, by using an enzyme preparation derived from Pseudomonas sp., the (91 ,10S)-enantiomer was hydrolyzed in a frans-specific fashion (i.e. via inversion of configuration at C-10) yielding the 9R,10R)-threo-diol. The remaining (9S,101 )-epoxide was converted into (-1-)-dispar lure, the sex pheromone of the gypsy moth in >95% ee [101]. 

This enzyme [EC 2.6.1.19] catalyzes the reversible reaction of y-aminobutyrate with a-ketoglutarate to yield succinate semialdehyde and glutamate. A number of enzyme preparations have been reported to also use )3-alanine, 5-aminopentanoate, and (i ,5)-3-amino-2-methylpropanoate as substrates. 

The use of enzymes as digestive aids is only applied under specific medical circumstances. Some medical conditions (e.g. cystic fibrosis) can result in compromised digestive function due to insufficient production/secretion of endogenous digestive enzymes. Digestive enzyme preparations are often formulated in powder (particularly tablet) form, and are recommended to be taken orally immediately prior to or during meals. As the product never enters the blood... 

An alternative to the synthesis of proteins by classical fragment synthesis in solution or by solid-phase synthesis on a support is the use of enzyme-catalyzed condensation of amino acids or peptides. This possibility was first demonstrated in 1938 91 with the synthesis of poorly soluble benzoyl-leucyl-leucine anilide by papain catalysis. After many years, this approach was extended to the preparation of peptide hormones such as Leu-enkephalin 92 and dynorphin(l -8).[93 This was made possible by the use of highly purified enzymes and by careful control of reaction conditions. The basic principles of protease-catalyzed peptide bond formation have been discussed.194 ... 

The use of enzymes for the manufacture of leather played a major role for the industrial scale production of enzymes. For the preparation of hides and skins for tanning, the early tanners kept the dehaired skins in a warm suspension of the dungs of dogs of birds. Wood was the first in 1898 to show that the bating action of the unpleasant dungs was caused by the enzymes (pepsin, trypsin, lipase) which they contained. In the context... 

Once the appropriate starting compound has been selected and the number of routes to the product has been reduced, questions concerning the biocatalyst arise. It largely depends on the reaction that has to be catalyzed whether it is possible to use cell-free extracts, or whether it is necessary to use purified enzyme preparations (see paragraph 5.6) or even growing or resting whole cells. Some of the criteria that play a role in deciding what catalyst to choose have been listed in Table 5.4. 

Figure 9.2 A schematic presentation of different types of enzyme preparations used in non conventional media, a enzyme powder, b enzyme crystals c enzyme on a porous support d covalently modified enzyme dissolved in the solvent e enzyme solubilised by surfactant ...

The N-acetylneuraminic acid derivative 44 is widely distributed. It was isolated from a strain182 of Escherichia coli, and has been obtained from cytidine 5 -triphosphate and N-acetylneuraminic acid by the action of enzyme preparations from Neisseria meningitidis183 and from animal tissues.184-186 The latter enzyme can also make use of N-glycolylneuraminic acid as a substrate, to give the respective cytidine 5 -phosphate derivative. 

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