1 research outputs found
Frontal analysis for characterizing the adsorption-desorption behavior of beta-lactoglobulin on immunoadsorbents
High-performance frontal affinity chromatography was employed to study the adsorption–desorption kinetics characterizing the retention of
Beta-lactoglobulin (Beta-LG) onto polyclonal anti-Beta-lactoglobulin (anti-Beta-LG) chromatographic supports. The adsorption and desorption processes were
studied by analyzing two different elution fronts separated by a relatively long rinsing step. The method consists in performing two successive frontal
injections of the protein. In between, the column was rinsed with a given volume of mobile phase (buffer alone). During this rinsing stage, a partial
desorption may occur and a novel amount of protein could be adsorbed in the second frontal injection step. The whole process (first adsorption,
possible desorption, and second adsorption) was simulated by a numerical procedure, in which the column was divided into a large number of slices.
A model based on bi-Langmuir type kinetics was used to describe the adsorption of the protein on the support. The model assumes a non-uniform
adsorbent with two types of binding sites. At equilibrium the adsorption isotherm is of the bi-Langmuir type. A global adsorption effect was
considered which includes the effective binding process and the mass transfer resistances due to the transport to the binding site. Therefore, the
column capacity and the kinetic parameters of the model (apparent adsorption and desorption rate constants) were determined from the best fit of
the first and second adsorption fronts to the experimental ones. The other parameters of the model are the saturation capacities for the adsorption on
each type of sites. The equilibrium affinity constants were estimated in a single experiment from the ratio of the apparent adsorption and desorption
rate constants. The high values found (around 108 M−1) reveal a strong interaction of -LG with the immunoadsorbent. Kinetic measurements
were carried out at different flow rates. Both the apparent adsorption and desorption kinetics were faster at larger flow rates, indicating an important
contribution of the mass transfer resistance in the stagnant fluid at the particle boundary. However, as expected, close values were found for the
resulting equilibrium constants calculated from the ratio of the apparent adsorption and desorption rate constant determined at various flow rates.A.P. acknowledges Spanish Ministry of Science and Tech-
nology for a predoctoral grant. This work has been supported
by Spanish CICYT (Project TIC2003-01906) and Foundation
Ramon Areces. Collaboration between laboratories has been
possible thanks to French-Spanish Cooperation between CNRS
and CSIC.Peer reviewe