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A low perfusion rate microreactor for continuous monitoring of enzyme characteristics: application to glucose oxidase.

Posthuma-Trumpie GA, Venema K, van Berkel WJ, Korf J - Anal Bioanal Chem (2007)

Bottom Line: The microreactor was used for on-line preparation of apoglucose oxidase in strong acid and its subsequent reactivation with flavin adenine dinucleotide.A low backpressure at very low flow rates is an advantage, which increases the sensitivity.A variety of further applications of the microreactor are suggested.

View Article: PubMed Central - PubMed

Affiliation: University of Groningen, University Medical Centre Groningen (UMCG), Department Psychiatry, Hanzeplein 1, 9713 EZ, Groningen, The Netherlands. truus.posthuma@wur.nl

ABSTRACT
This report describes a versatile and robust microreactor for bioactive proteins physically immobilized on a polyether sulfone filter. The potential of the reactor is illustrated with glucose oxidase immobilized on a filter with a cut-off value of 30 kDa. A flow-injection system was used to deliver the reactants and the device was linked on-line to an electrochemical detector. The microreactor was used for on-line preparation of apoglucose oxidase in strong acid and its subsequent reactivation with flavin adenine dinucleotide. In addition we describe a miniaturized version of the microreactor used to assess several characteristics of femtomole to attomole amounts of glucose oxidase. A low negative potential over the electrodes was used when ferrocene was the mediator in combination with horseradish peroxidase, ensuring the absence of oxidation of electro-active compounds in biological fluids. A low backpressure at very low flow rates is an advantage, which increases the sensitivity. A variety of further applications of the microreactor are suggested.

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Schematic design of the reaction compartment of the microreactor. (a) sketch; (b) schematic diagram. Shown here is the microreactor with one PES ultrafilter which is then connected to the flow system using FST and finger-tight fittings. The mesh screen is used to support the ultrafilter
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Fig1: Schematic design of the reaction compartment of the microreactor. (a) sketch; (b) schematic diagram. Shown here is the microreactor with one PES ultrafilter which is then connected to the flow system using FST and finger-tight fittings. The mesh screen is used to support the ultrafilter

Mentions: The overall design of the microreactors is shown in Fig. 1. They were micro machined from Delrin by the instrumental workshop of Groningen University. For preparation of apoGOx and reconstitution of GOx activity, a microreactor as described in Ref. [12] was used. In contrast to the original device, only one PES filter with a cut-off value of 30 kDa (Sartorius, Göttingen, Germany) and punched to a diameter of 13 mm, was used. Performance studies were done with our miniaturized design. The dimensions of the miniaturized microreactor were: outer length 3 cm, outer diameter 1.5 cm. A PES membrane filter was punched to a diameter of 4 mm and placed in the microreactor (effective diameter 3 mm, effective volume 1 μL) and connected between the injection valve and the detector with fused-silica tubing (FST) (150 μm OD, 50 μm ID) (Polymicro Technologies, Phoenix, AZ, USA) pinched off in Teflon tubing 1/16 inch OD and 170 μm ID (Aurora Borealis Control, Schoonebeek, The Netherlands) with use of finger-tight fittings, to avoid high dead volumes.Fig. 1


A low perfusion rate microreactor for continuous monitoring of enzyme characteristics: application to glucose oxidase.

Posthuma-Trumpie GA, Venema K, van Berkel WJ, Korf J - Anal Bioanal Chem (2007)

Schematic design of the reaction compartment of the microreactor. (a) sketch; (b) schematic diagram. Shown here is the microreactor with one PES ultrafilter which is then connected to the flow system using FST and finger-tight fittings. The mesh screen is used to support the ultrafilter
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2117337&req=5

Fig1: Schematic design of the reaction compartment of the microreactor. (a) sketch; (b) schematic diagram. Shown here is the microreactor with one PES ultrafilter which is then connected to the flow system using FST and finger-tight fittings. The mesh screen is used to support the ultrafilter
Mentions: The overall design of the microreactors is shown in Fig. 1. They were micro machined from Delrin by the instrumental workshop of Groningen University. For preparation of apoGOx and reconstitution of GOx activity, a microreactor as described in Ref. [12] was used. In contrast to the original device, only one PES filter with a cut-off value of 30 kDa (Sartorius, Göttingen, Germany) and punched to a diameter of 13 mm, was used. Performance studies were done with our miniaturized design. The dimensions of the miniaturized microreactor were: outer length 3 cm, outer diameter 1.5 cm. A PES membrane filter was punched to a diameter of 4 mm and placed in the microreactor (effective diameter 3 mm, effective volume 1 μL) and connected between the injection valve and the detector with fused-silica tubing (FST) (150 μm OD, 50 μm ID) (Polymicro Technologies, Phoenix, AZ, USA) pinched off in Teflon tubing 1/16 inch OD and 170 μm ID (Aurora Borealis Control, Schoonebeek, The Netherlands) with use of finger-tight fittings, to avoid high dead volumes.Fig. 1

Bottom Line: The microreactor was used for on-line preparation of apoglucose oxidase in strong acid and its subsequent reactivation with flavin adenine dinucleotide.A low backpressure at very low flow rates is an advantage, which increases the sensitivity.A variety of further applications of the microreactor are suggested.

View Article: PubMed Central - PubMed

Affiliation: University of Groningen, University Medical Centre Groningen (UMCG), Department Psychiatry, Hanzeplein 1, 9713 EZ, Groningen, The Netherlands. truus.posthuma@wur.nl

ABSTRACT
This report describes a versatile and robust microreactor for bioactive proteins physically immobilized on a polyether sulfone filter. The potential of the reactor is illustrated with glucose oxidase immobilized on a filter with a cut-off value of 30 kDa. A flow-injection system was used to deliver the reactants and the device was linked on-line to an electrochemical detector. The microreactor was used for on-line preparation of apoglucose oxidase in strong acid and its subsequent reactivation with flavin adenine dinucleotide. In addition we describe a miniaturized version of the microreactor used to assess several characteristics of femtomole to attomole amounts of glucose oxidase. A low negative potential over the electrodes was used when ferrocene was the mediator in combination with horseradish peroxidase, ensuring the absence of oxidation of electro-active compounds in biological fluids. A low backpressure at very low flow rates is an advantage, which increases the sensitivity. A variety of further applications of the microreactor are suggested.

Show MeSH
Related in: MedlinePlus