Monolithic bioreactors: Effect of chymotrypsin immobilization on its biocatalytic properties

Ponomareva, E. A.; Kartuzova, V.E.; Влах, Е. Г.; Tennikova, Tatiana

doi:10.1016/j.jchromb.2010.01.001

Cited by 32 publications

(28 citation statements)

References 41 publications

(40 reference statements)

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“…The effect of flow rate on the activity of methacrylatebased ultrashort monolithic minicolumn with a diameter of 5.2 mm and a thickness of 5.0 mm (CIM disc) carrying CT was first studied by Tennikova's group [28]. In their study, a similar behaviour for the effect of flow rate on the conversion obtained in the monolithic minireactor was also observed.…”

Section: Resultsmentioning

confidence: 85%

“…Different micro-IMERs carrying CT, acetylcholinesterase, β-secretase or pectin lyase were fabricated using CIM disc as solid matrix [10,20,28]. All these IMERs were operated with the mobile phase flow rates in the range of 0.2-1.5 mL/min and by injecting sample substrate solutions with volumes ranging between 10 and 50 μL.…”

Section: Resultsmentioning

confidence: 99%

“…On the other hand, the IMER containing immobilised CT onto a CIM disc was operated in a different form, by circulating constant volume of substrate solution for a certain period with the flow rates in the range of 0.2-1.5 mL/min [28]. In the case of our system, satisfactory substrate conversions higher than 60% (w/w) were achieved by continuous feeding of substrate solution with reasonably low flow rates in the range of 25-60 μL/min.…”

Section: Resultsmentioning

confidence: 99%

“…In these studies, mostly silica-based monoliths were utilised as the support matrix [26][27][28][29]. In present study, by considering the common use of CT in monolithic bioreactors, a new IMER in the form of "monolithic micro-column" was prepared by immobilisation of CT on a polyacrylate-based monolith via click-chemistry.…”

Section: Introductionmentioning

confidence: 99%

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Synthesis of a monolithic, micro-immobilised enzyme reactor via click-chemistry

2012

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An immobilised enzyme reactor (IMER) in the form of capillary monolith was developed for a micro-liquid chromatography system. The plain monolith was obtained by in situ thermal copolymerisation of glycidyl methacrylate and ethylene dimethacrylate in a fused silica capillary (200 × 0.53 mm ID) by using n-propanol/1,4-butanediol as porogen. The enzyme, α-chymotrypsin (CT), was covalently attached onto the monolith via triazole ring formation by click-chemistry. For this purpose, the monolithic support was treated with sodium azide and reacted with the alkyne carrying enzyme derivative. CT was covalently linked to the monolith by triazole-ring formation. The activity behaviour of monolithic IMER was investigated in a micro-liquid chromatography system by using benzoyl-L-tyrosine ethyl ester (BTEE) as synthetic substrate. The effects of mobile-phase flow rate and substrate feed concentration on the final BTEE conversion were investigated under steady-state conditions. In the case of monolithic IMER, the final substrate conversion increased with increasing feed flow rate and increasing substrate feed concentration. Unusual behaviour was explained by the presence of convective diffusion in the macropores of monolith. The results indicated that the monolithic-capillary IMER proposed for micro-liquid chromatography had significant advantages with respect to particle-based conventional high-performance liquid chromatography-IMERs.

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Section: Resultsmentioning

confidence: 85%

Section: Resultsmentioning

confidence: 99%

Section: Resultsmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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Synthesis of a monolithic, micro-immobilised enzyme reactor via click-chemistry

2012

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“…Polymer aldehyde groups formed after activation allow fast covalent binding of amino group bearing ligands under mild conditions, with elimination of water as the only side product (Ponomareva et al, 2010). The glutaraldehyde spacer (Fig.…”

Section: Immobilization On a Cim Disk With Spacer Armmentioning

confidence: 99%

The Value of Fungal Protease Inhibitors in Affinity Chromatography

Sabotič¹,

Koruza²,

Gabor³

et al. 2012

Affinity Chromatography

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Miniaturized biocatalysis on polyacrylate‐based capillary monoliths

Elsner

Ernst

Buchmeiser

2010

J of Applied Polymer Sci

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We developed a synthetic concept for the immobilization of enzymes onto monolithic supports. In addition, we elaborate on a generally applicable method for the rapid screening of the activity of such immobilized enzymes. For these purposes, we prepared monolithic acrylate-based systems by electron-beam (EB)-triggered freeradical polymerization within the confines of 200-lm capillary columns. Aiming for protein immobilization, we subjected the polyacrylate-based monoliths to EB-mediated grafting processes to introduce functional surface-located groups suited for the subsequent generation of functional units that themselves could bind different proteins. For the generation of the functional units, we used ring-opening metathesis polymerization and free-radical polymerization. The produced systems were tested for their ability to bind or repel proteins as exemplified by the use of the serine protease trypsin, which was used to catalyze the hydrolysis of N-a-benzoyl-L-arginine ethyl ester (BAEE). Finally, the monolith-immobilized trypsin systems were used for enzymatic peptide synthesis purposes, such as the acyl transfer of BAEE to amino acid amides. Complementarily, we used an immobilized trypsin variant, which we additionally subjected to on-column chemical modification with succinic anhydride to alter its synthetic properties.

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Monolithic bioreactors: Effect of chymotrypsin immobilization on its biocatalytic properties

Cited by 32 publications

References 41 publications

Synthesis of a monolithic, micro-immobilised enzyme reactor via click-chemistry

Synthesis of a monolithic, micro-immobilised enzyme reactor via click-chemistry

The Value of Fungal Protease Inhibitors in Affinity Chromatography

Miniaturized biocatalysis on polyacrylate‐based capillary monoliths

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