Unexpected differences in the behavior of ovotransferrin at the air–water interface at pH 6.5 and 8.0 Cécile Le Floch-Fouéré a,b , Stéphane Pezennec a,b,⇑ , Michel Pézolet c , Jean-François Rioux-Dubé c , Anne Renault d , Sylvie Beaufils d a INRA, UMR1253 Science et Technologie du Lait et de l’Œuf, F-35042 Rennes, France b AGROCAMPUS OUEST, UMR1253 Science et Technologie du Lait et de l’Œuf, F-35042 Rennes, France c Centre de recherche sur les matériaux avancés, Département de Chimie, Université Laval, Québec, Canada G1V 0A6 d Institut de Physique de Rennes, UMR6251 UR1-CNRS, Université de Rennes 1, F-35042 Rennes, France article info Article history: Received 28 October 2010 Accepted 21 January 2011 Available online 27 January 2011 Keywords: Protein Adsorption Air–water interface Conformation Ovotransferrin Charge abstract Adsorption of purified apo-ovotransferrin at the air–water interface was studied by ellipsometry, surface tension, polarization–modulation infrared reflection–absorption spectroscopy (PM-IRRAS), and shear elastic constant measurements. No significant difference was observed between pH 6.5 and 8.0 as regards the final value of surface concentration and surface pressure. However at low concentration, a weak bar- rier to adsorption is evidenced at pH 6.5 and confirmed by PM-IRRAS measurements. At a pH where the protein net charge is negative (pH 8.0), the behavior of ovotransferrin at the air–water interface is more influenced by charge effects rather than bulk concentration effects. At this pH, the interface exhibits a low shear elastic constant and a spectral signature not usual for globular proteins. Ó 2011 Elsevier Inc. All rights reserved. 1. Introduction Ovotransferrin, also called conalbumin, constitutes about 13% of egg-white proteins [1]. It is a member of the transferrins, soluble glycoproteins implicated in the regulation and transport of iron in vertebrates [2]. It is a protein with an iso-electric point (pI) of 6.5, of known sequence and three dimensional structure [3,4]. Its amino acid sequence consists of a single chain of 686 residues (77.7 kDa) organized in two lobes of similar conformation (lobe N and lobe C) linked together with a short connecting peptide. Each lobe is made of two domains containing a single high-affinity iron- binding site in the inter-domain cleft, enabling ovotransferrin to exhibit antimicrobial activity through iron deprivation [5,6]. While most studies on ovotransferrin have been devoted to the understanding of the mechanism of uptake and release of metallic ions, notably ferric ions, little experiments were dedicated to techno-functional properties of this protein. The analysis of the gelation properties of apo-ovotransferrin by DSC has allowed characterizing its thermal denaturation around 65 °C at pH 7.0: it is the most heat-sensitive protein of egg white [7–9]. Its foaming properties seem intermediate between those of ovalbumin and lysozyme [10,11]. Recent advances on the relationship between molecular, interfacial and foaming properties, may lead to a better understanding of foaming properties of food proteins [12–15]. Numerous data are available about the kinetics and thermodynam- ics of protein adsorption, as provided by surface tension measure- ments, ellipsometry and radiolabelling techniques [16–21]. The interfacial behavior of globular proteins (lysozyme, b-lactoglobu- lin, bovine serum albumin, ovalbumin) has been extensively stud- ied at the air–water interface [17,21–26]. However the kinetics of ovotransferrin adsorption at the air–water interface and the influ- ence of bulk protein concentration on the kinetics have not been investigated in a specific way. Damodaran et al. studied the com- petitive adsorption of five major egg-white proteins, namely, lyso- zyme, ovalbumin, ovotransferrin, ovomucoid and ovoglobulins. In this case, it seems that ovotransferrin, lysozyme and ovomucoid are totally excluded from the interface [27]. Yet, results obtained about co-adsorption and foaming properties of mixed proteins demonstrate the potential dramatic impact of one minor compo- nent of the mixture [28]. Thus, with the long-term objective to understand interfacial properties of protein mixtures, it seems rel- evant to first characterize specifically the interfacial behavior of this protein, which is the second most abundant protein in hen egg white. Furthermore, understanding the biochemical and phys- ico-chemical determinants of protein interfacial behavior has to be enriched with data about scarcely characterized proteins. 0021-9797/$ - see front matter Ó 2011 Elsevier Inc. All rights reserved. doi:10.1016/j.jcis.2011.01.073 ⇑ Corresponding author at: INRA, UMR1253 Science et Technologie du Lait et de l’Œuf, 65 rue de Saint-Brieuc, 35042 Rennes CEDEX, France. E-mail address: stephane.pezennec@rennes.inra.fr (S. Pezennec). Journal of Colloid and Interface Science 356 (2011) 614–623 Contents lists available at ScienceDirect Journal of Colloid and Interface Science www.elsevier.com/locate/jcis