Encapsulation of Milk β-Lactoglobulin by Chitosan Nanoparticles

Naturally occurring polymers, such as chitosan, have been extensively studied as carriers for therapeutic protein and gene delivery systems. beta-Lactoglobulin (beta-LG) is a member of the lipocalin superfamily of transporters for small hydrophobic molecules. We examine the binding of milk beta-lactoglobulin with chitosan of different sizes such as chitosan 15, 100, and 200 KD in aqueous solution at pH 5-6, using FTIR, CD, and fluorescence spectroscopic methods. Structural analysis showed that chitosan binds beta-LG via both hydrophilic and hydrophobic contacts with overall binding constants of K beta-LG-ch-15 = 4.1 (+/- 0.4) x 10(2) M-1, K beta-LG-ch-100 = 7.2 (+/- 0.6) x 10(4) M-1, and K beta-LG-ch-200 = 3.9 (+/- 0.5) x 10(3) M-1 with the number of bound protein per chitosan (n) 0.9 for ch-15, 0.6 for ch-100, and 1.6 for ch-200. Chitosan 100 KD forms stronger complexes with beta-LG than chitosans 200 and 15 KD. Polymer binding did not alter protein conformation inducing structural stabilization. Chitosan 100 is a stronger protein transporter than chitosan 15 and 200 KD.