Immobilized p-galactosidase

Temperature explicit functions for inactivation parameters considering substrate and product modulation and a two-stage series mechanism were also determined and validated as already presented in Table 5.2. Temperature optimization of CPBR was then accomplished by replacing these temperature-explicit functions in Eqs. 5.76 and 5.79. A battery of staggered reactors was considered to absorb output flow fluctuations due to enzyme inactivation (see Eqs. 5.82 and 5.83). Elow-rate was considered constant during each time interval (see Eq. 5.84) so that variation in substrate conversion was below 1%. Conditions of operation and design parameters used in the simulation of an industrial CPBR for the hydrolysis of whey permeate with immobilized p-galactosidase are summarized in Table 5.4. 

Table 5.5 Temperature Optimization Based on a Cost-Objective Function of Multiple Staggered CPBER with Chitin-Immobilized P-Galactosidase Under Biocatalyst Replacement at 25% of Initial Activity. Operation Conditions Are Those in Table 5.3...

Except the LBL deposition for the enzyme immobilization, lactose has been hydrolyzed using covalently immobilized P-galactosidase on thermally stable carrageenan coated with chitosan. The immobilized enzyme showed lactose conversion of 70 % at 7 h compared to 87 % for the free enzyme. However, the... 

Brena BM, Irazoqui G, Giacomini C, Batista-Viera F. Effect of increasing co-solvent concentration on the stabihty of soluble and immobilized p-galactosidase. J Mol Catal B Enzym 2003 21 25-29. 

Ferric ion was immobilized on a Chelating Sepharose Fast Flow column preparatory to the separation of seven enkephalin-related phosphopep-tides.17 Non-phosphorylated peptides flowed through the column, and the bound fraction contained the product. The capacity of the column was found to be 23 pmol/mL by frontal elution analysis. Cupric ion was immobilized on Chelating Superose for the isolation of bovine serum albumin.18 Cupric ion was immobilized on a Pharmacia HiTrap column for the separation of Protein C from prothrombin, a separation that was used to model the subsequent apparently successful separation of Factor IX from prothrombin Factor IX activity of the eluate was, however, not checked.19 Imidazole was used as the displacement agent to recover p-galactosidase from unclarified homogenates injected onto a nickel-loaded IMAC column.20 Pretreatment with nucleases and cleaning in place between injections were required procedures. A sixfold purification factor was observed. 

Clemmit, R.H. and Chase, H.A., Immobilized metal affinity chromatography of P-galactosidase from unclarified Escherichia coli homogenates using expanded bed adsorption, /. Chromatogr. A, 874, 27, 2000. 

Bakken, A.P., Hill, C.G. and Amnndson, C.H. (1992) Hydrolysis of Lactose in Skim Milk by Immobilized Beta Galactosidase Bacillus-Circulans. Biotechnology and Bioengineering, 39, 408-417. 

H. Liu, H. Li, T. Ying, K. Sun, Y. Qin and D. Qi, Amperometric biosensor sensitive to glucose and lactose based on co-immobilization of ferrocene, glucose oxidase, p-galactosidase and mutarotase in /i-cyclodextrin polymer, Anal. Chim. Acta., 358(2) (1998) 137-144. 

De Rosa, M, Nicolaus, A., Gambacorta, B., Buonocore, V., and Poerio, E. (1980) Immobilized bacterial cells containing athermostable p-galactosidase, Biotechnol. Letters 2, 29 -34... 

Coupling proteins (by covalent methods) to magnetic supports is the most common approach (105). Various classes of proteins, such as lectins (110) and antibodies (111), and a large number of enzymes including invertase (112,113), papain (106, 114) a-chymotrypsin (114, 115), P-galactosidase (114), adenylate kinase (112), acetate kinase (112), and horseradish peroxidase (112), have been immobilized. In fact, such supports are now commercially available, for example, Enzacryl FEO-M (available from Aldrich Chemictd Company, USA) and Dyna (sold by Dynal, Norway). 

Ariga et al. [48] have investigated the behavior of the monolith reactor in which Echerichia coli with P-galactosidase or Saccharomyces cerevisiae was immobilized within a thin film of K-carragcenan gel deposited on the channel wall. The effects of mass transfer resistance and axial dispersion on the conversion were studied. Those authors found that the monolith reactor behaved like the plug-flow reactor. The residence-time distribution in this reactor was comparable to four ideally mixed tanks in series. The influence of gas evolution on liquid film resistance in the monolith reactor was also investigated. It was shown that at low superficial gas velocities, the gas bubble may adhere to the wall, which decreases the effective surface area available for the reaction. The authors concluded that the reactor was very effective in the reaction systems accompanied by gas evolution, such as fermentations. 

Enzymes modified with carbohydrates (neoglycoenzymes) can be used in cytochemistry as described above or in biochemical detection of lectins in solid-phase assays to gain greater sensitivity in analysis. For example, bacterial 3-galactosidase modified with p-aminophenyl a-D-mannopyranoside via amide linkage was useful in determination of Con A immobilized on plastic microtiter plates, and lactose-modified P-galactosidase was effective in histochemical detection of galactoside-specific lectins [63]. Other enzymes frequently used for these applications are alkaline phosphatase and horse radish peroxidase. There are a number of colorimetric, fluorometric, and chemiluminescent substrates available for these enzymes. 

P-Galactosidase. Lysosomal P-galactosidase can be purified from human liver or placenta with conventional methods (Meisler, 1972 Sloan, 1972 Miyatake and Suzuki, 1975) or with affinity chromatography on immobilized />-aminophenyl- or 6-aminohexyl-thio-3-D-galactoside (Miller et al., 1977 Lo et al., 1979). The most rapid and convenient procedure seems to be the two-step method of Miller et al. (1977), which is reported to give a more than 20,000-fold purification with 41% yield. The affinity gel is commercially available. 

Mosbach K, Guilford H, Ohlsson R et al. (1972) General Ugands in affinity chromatography. Cofactor-substrate elution of enzymes bound to the immobilized nucleotides adenosine 5 -monophosphate and nicotinamide-adenine dinucleotide. Biochem J 127(4) 625-631 Moses V, Prevost C (1966) Catabolite repression of P-galactosidase synthesis in Escherichia coli. Biochem J 100(2) 336-353... 

P-Galactosidase Silanized magnetite with immobilized p-aminophenyl-h-D-thiogalactopyranoside [119]... 

ORIENTATION CONTROLLED IMMOBILIZATION STRATEGY FOR p-GALACTOSIDASE ON ALGINATE BEADS... 

M. S. Mohy Eldin, E. A. Hassan, M. R. Elaassar. P-Galactosidase Covalent Immobilization on the Surface of P-Benzoquinone-Activated Alginate Beads as a Strategy For Overcoming Diffusion Limitation. III.Effect of Beads Formulation Conditions on the Kinetic Parameters, The 1st International Conference of Chemical Industries Research Division 6-8 December, 2004. 

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