Applied Biochemistry and Biotechnology

, Volume 158, Issue 3, pp 747–760

A Novel Reversible pH-Triggered Release Immobilized Enzyme System

Article

DOI: 10.1007/s12010-008-8373-2

Cite this article as:
Gai, L. & Wu, D. Appl Biochem Biotechnol (2009) 158: 747. doi:10.1007/s12010-008-8373-2

Abstract

A novel immobilized enzyme system supported by poly(acrylic acid/N,N’-methylene-bisacryl-amide) hydrogel microspheres was prepared. This system exhibited characteristics of reversible pH-triggered release. The morphology, size, and chemical structure were examined through optical microscopy, particle size analyzer, and Fourier transform infrared spectrometer. Immobilization and release features were further investigated under different conditions, including pH, time, and microsphere quantity. Results showed the microspheres were regularly spherical with 3.8 ~ 6.6 μm diameter. Loading efficiencies of bovine serum albumin immobilized by gel entrapment and adsorption methods were 93.9% and 56.2%, respectively. The pH-triggered protein release of the system occurred when medium pH was above 6.0, while it was hardly detected when medium pH was below 6.0. Release efficiencies of entrapped and adsorbed protein were 6.38% and 95.0%, respectively. Hence, adsorption method was used to immobilize trypsin. Loading efficiency of 77.2% was achieved at pH 4.0 in 1 h. Release efficiency of 91.6% was obtained under optimum pH catalysis condition set at 8.0 and trypsin was free in solutions with retention activity of 63.3%. And 51.5% of released trypsin could be reloaded in 10 min. The results indicate this kind of immobilized enzyme system offers a promising alternative for enzyme recovery in biotechnology.

Keywords

pH-dependent hydrogelReversible pH-triggered releaseBSAp(AAc/MBA) microspheresImmobilized trypsin

Abbreviations

AAc

acrylic acid

MBA

N,N′-methylene-bisacrylamide

BSA

bovine serum albumin

TEMED

N,N,N′,N′-tetramethylethylenediamine

BAEE

N-Benzoyl-L-arginine ethylester hydrochloride

p(AAc/MBA)

poly(acrylic acid/N,N′-methylene-bisacryl-amide)

DpH

main particle size of p(AAc/MBA) microspheres at certain pH

ηL

loading efficiency

ηR

release efficiency

ηU

recovery efficiency

Ci

initial BSA concentration introduced

Cf

final BSA concentration in supernatants after immobilization

Cr

BSA concentration in supernatants after release

Cu

BSA concentration in supernatants after recovery

Vl

loading solution volume

Vr

release solution volume

Copyright information

© Humana Press 2008

Authors and Affiliations

  1. 1.Key Laboratory of Biomedical Information Engineering of Education Ministry, School of Life Science and TechnologyXi’an Jiaotong UniversityXi’anChina