Colloids and Surfaces B: Biointerfaces 46 (2005) 152–161 Effect of NaCl and peptide concentration on the self-assembly of an ionic-complementary peptide EAK16-II Yooseong Hong, Mark D. Pritzker, Raymond L. Legge, P. Chen ∗ Department of Chemical Engineering, University of Waterloo, 200 University Ave. W., Waterloo, Ont., Canada N2L 3G1 Received 26 June 2005; received in revised form 30 October 2005; accepted 1 November 2005 Abstract Previous work has examined the effects of such factors as pH and peptide concentration on the self-assembly of ionic-complementary peptides. This work focused on the effect of sodium chloride on the molecular self-assembly of an ionic-complementary peptide EAK16-II (AEAEAKAKAEAEAKAK). Surface tensions and dimensions of the self-assembled nanostructures were determined for a wide range of peptide and sodium chloride concentrations using axisymmetric drop shape analysis-profile (ADSA-P) and atomic force microscopy (AFM), respectively. The critical aggregation concentration, or critical self-assembly concentration (CSAC), of EAK16-II was not significantly affected by the presence of NaCl. However, the analysis of size variations in self-assembled nanostructures in response to changes in NaCl concentration indicated that the presence of NaCl does influence the resulting dimensions of the peptide nanostructures when the peptide concentration is below its CSAC. A critical NaCl concentration was identified at ∼20 mM, below which the equivalent radius of the peptide fibrils increased with increasing salt concentration, and above which the opposite response was observed. This critical NaCl concentration was further confirmed in the surface tension measurements, where the equilibrium surface tension and induction time of the peptide at low concentrations (<CSAC) decreased with increasing NaCl concentration up to approximately 20 mM and a further increase caused the opposite trend. © 2005 Elsevier B.V. All rights reserved. Keywords: Self-assembly; Peptides; Atomic force microscopy (AFM); Axisymmetric drop shape analysis-profile (ADSA-P); NaCl 1. Introduction An understanding of the process of peptide self-assembly has a wide variety of biomedical implications, including scaffolding in tissue engineering [1–3], controlled drug delivery [4,5] and surface bioengineering [6–11]. However, the practical applica- tion of peptide self-assembly has been hampered due to a lack of control methods for the self-assembly process, which is due largely to a poor understanding of the molecular mechanisms involved. In order to understand the peptide self-assembly mechanism, it is important to understand the factors that affect the molecular behavior of the peptides in solution. The physicochemical fac- tors include: (i) amino acid sequence; (ii) concentration of the peptide; (iii) molecular size; (iv) pH; (v) ionic strength; (vi) tem- perature; (vii) solvent composition; and (viii) substrate on which ∗ Corresponding author. Tel.: +1 519 888 4567x5586; fax: +1 519 746 4979. E-mail address: p4chen@cape.uwaterloo.ca (P. Chen). self-assembly forms. Of these factors, amino acid sequence, pH [12], and peptide concentration [13,14] have been investigated for a number of ionic-complementary peptides, including EAK16-I (AEAKAEAKAEAKAEAK, - + - + - + - +), EAK16-II (AEAEAKAKAEAEAKAK, -- ++ -- + +) and EAK16-IV (AEAEAEAEAKAKAKAK, ---- + + + +). Fung et al. [14] and Hong et al. [13] studied the effect of peptide concentration on its self-assembly and found that there is a critical self-assembly concentration (CSAC) for EAK16-I and EAK16-II. An analogy to the CSAC can be made to the critical micelle concentration (CMC) for amphiphilic molecules such as surfactants. Amphiphilic surfactant molecules form micelles to minimize unfavorable interactions of hydrophobic tails with the surrounding water at or above the CMC. An amphiphilic peptide, with hydrophilic and hydrophobic amino acid residues, may also display similar concentration-dependent self-assembly behav- ior, leading to the formation of peptide nanostructures [12]. For a series of ionic-complementary peptides (EAK16s), Hong et al. [12] found that the self-assembly process is depen- dent on the charge distribution along the peptide backbone. 0927-7765/$ – see front matter © 2005 Elsevier B.V. All rights reserved. doi:10.1016/j.colsurfb.2005.11.004