Biochemical Engineering Journal 14 (2003) 27–36 Determination of apparent kinetic parameters for competitive product inhibition in packed-bed immobilized enzyme reactors Ahmet R. Özdural a,∗ , Deniz Tanyolaç a , ˙ Ismail H. Boyacı b , Mehmet Mutlu b , Colin Webb c a Chemical Engineering Department, Hacettepe University, Beytepe, 06532 Ankara, Turkey b Food Engineering Department, Hacettepe University, Beytepe, 06532 Ankara, Turkey c Department of Chemical Engineering, Satake Centre for Grain Process Engineering, UMIST, P.O. Box 88, Manchester M60 1QD, UK Received 14 March 2002; accepted after revision 25 July 2002 Abstract In this study, a simple and effective technique for characterizing Michaelis–Menten type kinetics with competitive product inhibition in packed-bed re-circulated immobilized enzyme reactors is presented, where the use of nonlinear regression techniques for multi-parameter estimation are not required. In order to demonstrate the new technique introduced in this work, enzymatic conversion of lactose in a recycling packed-bed reactor is envisaged where -galactosidase (lactase, EC 3.2.1.23) enzyme is immobilized on a weak base ion exchanger resin (Duolite A 568). For the experimental conditions used in this research, the total competitive inhibition by product (galactose) model is sufficient to represent the lactose hydrolysis kinetics in a packed-bed reactor. © 2002 Elsevier Science B.V. All rights reserved. Keywords: Competitive inhibition; Immobilized enzymes; Kinetic parameters; Lactose hydrolysis; Modeling; Packed-bed bioreactors 1. Introduction Enzymes are often used in an immobilized form in in- dustry because immobilized enzyme processes may be per- formed continuously and offer the possibility of reutilizing the enzyme. Other benefits obtained as well, include better operational control, flexibility of reactor design, and ease of product recovery without catalyst contamination [1]. The ki- netic parameters for the immobilized enzyme may be differ- ent from those of the soluble enzyme because of, amongst others, diffusion restrictions and interactions with the car- rier or due to immobilization. The value of the kinetic pa- rameters of the immobilized biocatalyst, in the presence of above-mentioned factors, is referred to as apparent kinetics. Furthermore, due to the difference in hydrodynamic condi- tions, the immobilized enzyme apparent kinetic parameters determined through batch wise stirred reactor experiments usually do not reflect the apparent kinetic parameters of immobilized enzymes in continuously operated packed-bed reactors. Under steady-state conditions and without inhibition the apparent kinetic parameters in packed-bed enzyme reactors ∗ Corresponding author. Tel.: +90-312-297-7475; fax: +90-312-299-2124. E-mail address: ozdural@hacettepe.edu.tr (A.R. Özdural). are usually calculated from a κ S in versus ln(1 - κ) plot with the following equation [2]: FκS in = K ′ m F ln(1 - κ) + v ′ max V eff (1) Özdural et al. [3] proposed a new technique for the deter- mination of kinetic parameters for packed-bed immobilized enzyme reactions under unsteady-state conditions if inhibi- tion is negligible. The techniques for determining inhibition and Michaelis– Menten parameters of soluble enzymes are well known [4,5]. Inhibition of immobilized enzymes is an important topic and since the design equations of continuous enzyme reactors generally require the use of apparent kinetic parameters [6], these parameters must be known. Only nonlinear regression techniques are reported in the literature, for the determina- tion of immobilized enzyme inhibition kinetics in contin- uous reactors [7]. Nonlinear regression techniques usually require a large number of data points to fit. Furthermore, the outcome is valid only within the ranges of the evaluated data, otherwise it easily leads to erroneous conclusions. The present study was undertaken with the aim of devel- oping a new method for the retrieval of apparent parameters for competitive inhibition by product kinetics in packed-bed immobilized enzyme reactors, which does not rely on non- linear regression analyses, and is thus free from the proba- ble limitations of such techniques. Other types of inhibition 1369-703X/02/$ – see front matter © 2002 Elsevier Science B.V. All rights reserved. PII:S1369-703X(02)00099-2