Controlled Pore Silica Research Articles

Stabilization and reutilization ofBacillus megaterium glucose dehydrogenase by immobilization

Glucose dehydrogenase (GDH) from Bacillus megaterium was immobilized using aminopropyl controlled-pore silica (CPS, average pore sizes of 170 and 500 A) as a support and glutaraldehyde as a bifunctional crosslinking agent. The CPS-immobilized enzyme could be reused 12 times and the best results were obtained using aminopropyl CPS-500 and bovine serum albumin as a feeder for stabilizing the protein layer on the support. DEAE-Sephadex (A-25 and A-50) was also used as a support for immobilizing GDH, with yields of around 42% for A-25 and 25-30% for A-50. The effect of pH on the immobilization procedure showed pH 6.5 to be better than pH 7.5 with respect to the recovery of enzyme activity. Both preparations of DEAE-Sephadex immobilized GDH could be reused several times and were thermostable at 40 degrees C for 7 h. The kinetic parameters as Michaelis constant and maximum rate were determined for the immobilized enzyme and compared with those for the freeform.

Difructose anhydride-forming bacterial inulinase II and fructogenic fungal inulinase I

The reserve polymeric inulin from dahlia tuber (>12% or >60% yield, wet or dry basis, respectively) follows as an attractive source for both free fructose or difructofuranose anhydride (DFA III). Although DFA biological activity is not completely understood, there is interest in characterizing other DFA III-producers besides Arthrobacter ureafaciens. The inulinolytic bacterial isolate named YLW, owing to the yellow hue in agar slants, is such a producer. Its biochemical characterization showed the presence of galactosylated and mannosylated glycolipids associated with the bacterial cells. Immobilization of fungal inulinase I and bacterial inulinase II, the respective enzymatic catalysts for the production of fructose and DFA III by inulin hydrolysis, was attempted using controlled-pore silica (CPS). The effects of pH, temperature, and incubation time was analyzed and compared for both enzymes in the free and immobilized forms.

Performance of fixed and fluidized bed reactors with immobilized enzyme

Saccharification of α-amylase liquefied cassava starch was carried out at pH 4.5 and 45‡C, in both fixed and fluidized bed reactors, using amyloglucosidase immobilized in 0.5 mm controlled pore silica particles. Reactor performance data are compared with mathematical modeling. Data show that for equal normalized residence time and lower fluid-bed porosities, the fluidized bed mode leads to higher conversions than fixed bed. Interparticle mass transfer experiments showed absence of diffusion limitations and starch conversion reached 98.5% with a real residence time of only 10 min, whereas the conventional liquid-phase process requires 48 h to achieve the same conversion.

Axial dispersion in a liquid fluidized bed of particles akin to immobilized enzymes

The axial dispersion of a liquid fluidized bed of controlled pore silica (CPS) particles has been determined by the pulse tracer method. The CPS used was the same as for enzyme immobilization, having an average diameter of 0.436 mm and mean pore size of 37.5 nm. The fluidization liquid is α-amylase liquefied manioc starch, 30% w/v, 45°C pH=4.5. Nominal bed porosities tested were 0.7 and 0.8. The results show that the axial dispersion coefficient increases with greater superficial liquid velocities. Various available correlations tested disagree with each other to a large extent and are unable to represent collected experimental data.

Immobilization of glutamate decarboxylase and the preparation of an enzyme column for the synthesis of γ-[ 13N]aminobutyric acid

The enzyme glutamate decarboxylase ( l- glutamate 1-carboxy-lyase, EC 4.1.1.15) is used for the preparation of 1 μmol γ-[ 13 N] aminobutyric acid, a radiopharmaceutical designed for positron emission tomography. To obtain a rapid and high-yield synthesis of the labelled compound, glutamate decarboxylase was immobilized on controlled pore silica beads ( pore size 50 nm, particle size 100–200 μm), which were packed into a column. A screening of different immobilization techniques is described. The selected immobilization procedure was further optimized. The synthesis of γ-[ 13 N] aminobutyric acid as function of enzyme loading, column dimensions and flow rate was investigated.

Mechanism for reversible inactivation of immobilized ?-galactosidase from Bacillus circulans during continuous production of galacto-oligosaccharides

The specific activity-dependent stability of the immobilized β-galactosidase-2 (β-d-galactoside galactohydrolase, EC 3.2.1.23) from Bacillus circulans during the continuous production of galactooligosaccharides from lactose was studied. This was done by measuring the elution pattern of saccharides from the various immobilized Merckogel (controlled pore silica gel) columns and the amount of saccharides remaining in the gel. It was suggested that oligosaccharides produced were trapped inside the three dimensional enzyme aggregate with the immobilized enzyme having a specific activity of 240 units/g of wet gel, causing gradual inactivation, while the immobilized enzyme with 15 units/g of wet gel was stable since the oligosaccharides were not accumulated.

Continuous production of galacto-oligosaccharides from lactose using immobilized ?-galactosidase from Bacillus circulans

β-Galactosidase-2 (β-d-galactoside galactohydrolase, EC 3.2.1.23) from Bacillus circulans was purified using hydroxyapatite gel chromatography and immobilized onto Duolite ES-762 (phenolformaldehyde resin) and Merckogel (controlled pore silica gel) for continuous production of galacto-oligosaccharides using lactose as the substrate. The maximum amount of ologosaccharides produced by the immobilized enzyme was 35–40% of the total sugar during hydrolysis of 4.56% lactose. Partially purified β-galactosidase from B. circulans was also immobilized onto various supports for the same purpose. The stability of the immobilized β-galactosidase-2 or partially purified enzyme during a continuous reaction depended on their supports and specific activity. Of the supports tested, Merckogel was best for operational stability. With this support, the enzyme was quite stable with specific activity up to 15 units/g of wet gel; it was reversibly inactivated with more.

Enzymic interesterification of fats: Factors influencing the choice of support for immobilized lipase

Three grades of diatomaceous earth (Celite 560, Filtercel and Hyflo Supercel) and a controlled-pore silica have been examined for their suitability as support materials for lipase (triacyglycerol acylhydrolase, EC 3.1.1.3) catalysing the interesterification of fats. The controlled-pore silica gave a preparation with a low activity. Although all three Celites gave preparations with similar lipolytic activities, Hyflo Supercel gave the highest interesterification activity. The distribution of enzyme protein in Hyflo Supercel was examined by transmission electron microscopy.

Production of high glucose syrups using glucoamylase immobilized on alkylamine derivatives of titanium(IV)-activated porous silica: Kinetics and operat

Glucoamylase (EC 3.2.1.3.) was immobilized on controlled pore silica by a new covalent method of enzyme immobilization on inorganic supports. A 30 wt.%

Aromatic amine coupling ofaspergillus niger lactase to controlled-pore silica witho-dianisidine

Lactose (β-galactosidase) derived fromAspergillus niger was immobilized on controlledpore silica with an average pore diameter of 425 A. The coupling of this enzyme to the surface of the silica was accomplished by reacting the surface of the silica with o-dianisidine followed by the functionalization of the residual amine with glutaraldehyde or with nitrite to form the diazonium salt. The PH profiles of the immobilized enzymes were determined and compared. Continuous reactor studies of the glutaraldehyde-functionalized, immobilized enzyme indicated a half-life of 52 days at 50°C with a 5% lactose feed at pH 3.5.