Enzymes, immobilized derivatives

METALLOCENE DERIVATIVES RADIATION CROSSLINKING TCNQ DERIVATIVES ENZYME IMMOBILIZATION... 

It was reported that PEGylated lipase entrapped in PVA cryogel could be conveniently used in organic solvent biocatalysis [279], This method for enzyme immobilization is more convenient in comparison to other types of immobilization that take advantage of enzyme covalent linkage to insoluble matrix, since the chemical step which is time consuming and harmful to enzyme activity is avoided. The application of this catalytic system to the hydrolysis of acetoxycoumarins demonstrated the feasibility of proposed method in the hydrolysis products of pharmaceutical interest and to obtain regioselective enrichment of one of the two monodeacetylated derivatives. 

TCNQ derivatives Phthalocyanine derivatives Enzyme immobilization on surfaces Radiation crosslinking Metal carbonyl derivatives Organosilicon derivatives... 

Immobilized Derivatives of Cobalamins and Their Use as Affinity Adsorbents for a Study of Enzyme-Coenzyme Interaction... 

Free enzyme versus immobilized enzyme can influence the yield of lOS, additionally an immobilized system would be favorable economically as the biocatalyst can be reused, enables continuous production and the end product is free of contamination. Kim et al. [275] intended to make a comparison between the reaction kinetics of free and immobilized endo-inulinases in a batch reactor however significant differences were observed in the reaction behavior and product composition due to the form of enzyme used and the initial concentration of substrate. Yun et al. [276] investigated the effect of inulin concentration on the production of lOS by free and immobilized endo-inulinase from Pseudomonas sp. Their findings corroborate those of Kim et al. [275] whereby different products are formed depending on the form of enzyme a soluble enzyme yielded inulobiose and DP3 products, whereby the immobilized form predominantly produced inulobiose. As the concentration of inulin increased the yield of lOS did not increase in the soluble system and in the immobilized the yield remained the same. Although the enzyme was derived from Pseudomonas the immobilized form required a differ-... 

Foulds and Lowe (1986) combined mass production of the base sensor and enzyme immobilization as follows. Using gold or platinum ink, a working and counter electrode were deposited on a ceramic substrate. After thermal treatment of the electrode material a solution containing GOD and a pyrrole derivative of ferrocene was electrochemically polymerized at the electrode. The pyrrole component forms a conducting polymer and the immobilized ferrocene acts as electron acceptor for GOD. The structured immobilization permits this technique to be used for successive enzyme fixation to multiparameter sensors. 

When racemic aryl glycidyl ethers were subjected to aminolysis in aqueous buffer catalyzed by hepatic microsomal epoxide hydrolase from rat, the corresponding (S)-configurated amino-alcohols were obtained in 51-88% ee 131. On the other hand, when azide was employed as nucleophile for the asymmetric opening of 2-methyl-1,2-epoxyheptane in the presence of an immobilized crude enzyme preparation derived from Rhodococcus sp., which contains an epoxide hydrolase activity, the reaction revealed a complex picture 1321. The (S)-epoxide from the racemate was hydrolyzed (as in the absence of azide), and the less readily accepted (i )-enantiomer was transformed into the corresponding azido-alcohol (ee >60%). Although at present only speculations can be made about the actual mechanism of both the aminolysis and azidolysis reaction, in both cases it was proven that the reaction was catalyzed by a protein and that no reaction was observed in the absence of biocatalyst... 

Table 2 shows half-life time and inactivation coefficient for YLL soluble and immobilized on different supports. The enzyme immobilized on MANAE-agarose support presented lower stability than the soluble enzyme, perhaps because the immobilized derivative has been prepared in the presence of detergent to ensure the enzyme desegregation could be monomers, while soluble enzyme as dimers [27, 28]. Random immobilization may not really improve enzyme rigidity even in some cases, the enzyme stability may decrease after immobilization [10-14], e.g., if the support is able to establish undesired interactions with the enz)une. 

Immobilized enzyme stability was assayed by using 0.3 g of the immobilized CALB on fiber or 0.012 g of Novozyme 435 in successive batches of butyl butyrate synthesis. Assay conditions were the same as described for the determination of esterification yield. At the end of each batch, the immobilized lipase was removed from the reaction medium and rinsed with hexane (20 ml) to extract any substrate or product eventually retained in the matrix. After 1 h at room temperature, the immobilized derivative was introduced into a fresh medium. The residual conversion is given as percentage of initial conversion of butyric acid (first cycle of synthesis) under standard conditions (described in Esterification Yield Butyl Butyrate Synthesis ). 

Intraparticle diffusion can have a significant effect on the kinetic behavior of enzymes immobilized on solid carriers or entrapped in gels. In their basic analysis of this problem. Moo-Young and Kobayashi (1972) derived a general modulus and effectiveness factor. The results also predicted possible multiple steady-states as well as unstable situations for certain systems. While these results are very interesting it should be remembered that they are primarily mathematical and await extensive experimental support data. 

Roche D, Prasad K, Repic O (1999) Enantioselective acylation of 3-aminoesters using penicillin G acylase in organic solvents. Tetrahed Lett 40 3665-3668 Rolinson GN, Batchelor ER, Butterworth D et al. (1960) Formation of 6-aminopenicillanic acid from penicillin by enzymatic hydrolysis. Nat Lond 187 236-237 Rolinson GN, Geddes AM (2007) The 50th anniversary of the discovery of 6-aminopenicillanic acid (6-APA). Internatl J Antimicrob Agents 29 3-8 Resell CM, Ferndndez-Lafuente R, Guisdn JM (1993) Resolution of racemic mixtures by synthesis reactions catalyzed by immobilized derivatives of the enzyme peniciUin G acylase. J Mol Catal 84 365-371... 

Some results indicate that different attempts of FucA immobilization by covalent attachment provoked severe enzyme inactivation (Fessner and Walter 1997). FucA and DERA from E. coli and SHMT from Streptococcus thermophilus have been immobilized by multipoint covalent attachment to glyoxyl-agarose. Although this immobilization method had been very successful with many different enzymes (Guisdn et al. 1993), results obtained with these aldolases were dissimilar. For FucA, in spite of an immobilization yield of 80-90%, enzyme inactivation occurred during immobilization process and only 10-20% of activity was retained (Suau et al. 2005). On the other hand, SHMT immobilization yield was 100%, but the immobilized activity was lost during the sodium borohydride reduction step, probably due to the reduction of the Schiff base established between the cofactor (pyridoxal phosphate) and the aldolase. Finally, 100% of immobilization yield and 65% of retained activity in the immobilized derivative was achieved with DERA. 

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