Study of Structural Changes of Lactoferrin Using Flow Injection Analysis with Electrochemical Detection on Glassy Carbon Electrode

Ondrej Zitka,a,b Ales Horna,c Karel Stejskal,a,b Josef Zehnalek,a Vojtech Adam,a Ladislav Havel,d Ladislav Zeman,e Rene Kizeka,*

a Department of Chemistry and Biochemistry, d Department of Plant Biology, and
e Department of Animal Nutrition, Mendel University of Agriculture and Forestry,Zemedelska 1, CZ-613 00 Brno, Czech Republic.
Tel. +420(5)45133350,
Fax: +420(5)45212044,
E-mail: kizek@sci.muni.cz.

b
Department of Biochemistry, Faculty of Science, Masaryk University,Kotlarska 2, CZ-611 37 Brno, Czech Republic
c Department of Food Engineering and Chemistry, Faculty of Technology,Tomas Bata University, CZ-762 72 Zlin, Czech Republic

Paper based on a presentation at the 12th International Symposium on Separation Sciences, Lipica, Slovenia,
September 2729, 2006.

Abstract
Lactoferrin is considered to be a multifunctional protein. It appears to play several biologically important roles, where its structure is very crucial. The aim of this work was to investigate the basic electrochemical behaviour of lactoferrin by both stationary and flow electrochemical methods with respect its structural changes under various denaturing conditions. The electroactivity of lactoferrin was studied by three various electrochemical methods (linear sweep, differential pulse and square wave voltammetry). Based on the results obtained, we utilized flow injection analysis with electrochemical detection (FIA-ED) for determination of lactoferrin. We found out that the most suitable FIA-ED conditions were as follows: working electrode potential of 900 mV, Britton-Robinson buffer (pH 4.5) as the mobile phase, its flow rate 1 ml min1. Finally, we attempted to follow the changes of lactoferrin signal in the presence of chemical compounds or under the physical conditions leading to changes in its structure. As we have shown here, electrochemical analysis enables us to distinguish changes of protein structure easily and rapidly.

Keywords: lactoferrin, milk protein, linear sweep voltammetry, differential pulse voltammetry, square wave voltammetry, carbon paste electrode, glassy carbon electrode