Shing-Yi Suen

A mathematical analysis of affinity membrane bioseparations

Abstract A mathematical model including convection, diffusion and Langmuir adsorption is formulated to analyze the design and operation of affinity membrane bioseparations. A key prediction from the analysis is that the flow velocity through affinity membranes may be limited not by pressure drop, but rather by association kinetics between soluble protein and immobilized ligand. Furthermore, for thin membranes with Peclet numbers less than 40, slow association kinetics may limit the flow velocity to an extent where diffusion axial to flow undesirably becomes a dominant mechanism for transport of protein through the membrane. Lastly, even relatively small thickness variations of 3% and porosity variations of 1% may severely degrade membrane of performance. Practical consequences of these results are presented, e.g. constructing thicker membranes from stacks of thinner membranes will tend to reduce the effect of thickness and porosity variations, sharpen breakthrough curves, raise allowable flow velocities and increase loading capacities.

Analysis of protein adsorption on regenerated cellulose-based immobilized copper ion affinity membranes.

Immobilized metal affinity membranes (IMAMs) were prepared by immobilizing copper ions on microporous regenerated cellulose membranes through different types of chelating agents (dentate and triazine dye). The resulting chelator utilization percentages were 95% for iminodiacetic acid, 56% for N,N,N-tris(carboxymethyl)ethylenediamine, 52% for Cibacron blue 3GA, and 140% for Cibacron red 3BA. On the other hand, triazine dyes were slightly superior to dentate chelators on metal ion utilization for protein adsorption. In batch single-protein adsorptions, the protein adsorption capacity decreased with increasing molecular size and number of accessible surface histidine residues [lysozyme>bovine serum albumin(BSA)>gamma-globulin], while the binding strength order was the opposite (gamma-globulin>BSA>lysozyme). Moreover, the proportions of specific and nonspecific bindings were evaluated by varying pH and salt concentration conditions. A large fraction of the adsorption capacity was found to come from the nonspecific interactions for the prepared IMAMs. Lastly, batch three-protein adsorptions were performed and weak adsorption competition was observed.

Exploiting immobilized metal affinity membranes for the isolation or purification of therapeutically relevant species.

Increasing reports regarding the isolation or purification of biospecies for therapeutic purpose using the immobilized metal affinity chromatography have been presented in recent years. At the same time, membrane chromatography technique has also gained more and more attention for their advantage in speeding the separation process. The immobilized metal affinity membrane technique developed by combining these two techniques may provide an alternative potential tool for separating the therapeutically relevant biospecies. In this review paper, the features of the immobilized metal affinity membranes are discussed and concentrated on three subtopics: membrane matrices, immobilized metal affinity method, and membrane module designs. Several examples of practically applying the immobilized metal affinity membranes on the purification of potential therapeutics reported in the literature are subsequently presented. Lastly, this review also provides an overall evaluation on the possible advantages and problems existing in this technique to point out opportunities and further improvements for more applied development of the immobilized metal affinity membranes.

Sorption kinetics and breakthrough curves for pepsin and chymosin using pepstatin A affinity membranes

Isotherms and kinetic parameters for pepsin and chymosin sorption to immobilized pepstatin A were measured in batch experiments. The measured single-solute parameters were used in an affinity-membrane model which included competitive sorption kinetics, axial diffusion and dead volume mixing. The predictions made using the affinity-membrane model matched the experimental breakthrough curves, whereas predictions made using local-equilibrium theory were a distinct mismatch. The performance of affinity-membrane separations was dominated by slow sorption kinetics.

Single-Layered Graphene Oxide Nanosheet/Polyaniline Hybrids Fabricated Through Direct Molecular Exfoliation

In this study, we used direct molecular exfoliation for the rapid, facile, large-scale fabrication of single-layered graphene oxide nanosheets (GOSs). Using macromolecular polyaniline (PANI) as a layered space enlarger, we readily and rapidly synthesized individual GOSs at room temperature through the in situ polymerization of aniline on the 2D GOS platform. The chemically modified GOS platelets formed unique 2D-layered GOS/PANI hybrids, with the PANI nanorods embedded between the GO interlayers and extended over the GO surface. X-ray diffraction revealed that intergallery expansion occurred in the GO basal spacing after the PANI nanorods had anchored and grown onto the surface of the GO layer. Transparent folding GOSs were, therefore, observed in transmission electron microscopy images. GOS/PANI nanohybrids possessing high conductivities and large work functions have the potential for application as electrode materials in optoelectronic devices. Our dispersion/exfoliation methodology is a facile means of preparing individual GOS platelets with high throughput, potentially expanding the applicability of nanographene oxide materials.

A comparison of isotherm and kinetic models for binary-solute adsorption to affinity membranes

Single-solute isotherms for pepsin (EC 3.4.23.1) and chymosin (EC 3.4.23.4) adsorption to affinity membranes were fitted using five of the most popular isotherm models. It was found that the single-solute Langmuir isotherm was the best two-parameter model, although the three-parameter models gave even better fitting. Experimental binary-solute adsorption isotherms were compared with four different types of binary-solute Langmuir models using the single-solute parameters. The results showed that the difference in the saturation capacities affected the adsorption equilibrium. Furthermore, three types of binary-solute Langmuir models were converted into the kinetic form and used to calculate the association rate constants of pepsin and chymosin from experimental data. The best-fitted rate constant values were found to be identical for different kinetic models. However, the model predictions of association curves were significantly influenced when the values of association rate constants were changed.

Sorption kinetics and axial diffusion in binary-solute affinity-membrane bioseparations

Abstract A mathematical model, which includes convection, axial diffusion and Langmuir sorption kinetics, was developed which describes the separation performance of affinity membranes when two solutes are present. To illustrate the membrane performance, two affinity systems were analyzed: (1) two soluble monoclonal anitbodies (MAbs) binding to immobilized bovine serum albumin, where both MAbs had identical equilibrium binding constants but different sorption kinetics; and (2) soluble chymosin and pepsin binding to immobilized pepstatin A, where both enzymes had different equilibrium binding constants but nearly identical association kinetics. In the first system, the MAbs could not be separated based on local-equilibrium behavior, but could be cleanly separated based on differences in sorption kinetics. In the second system, the enzymes could be cleanly separated based on local-equilibrium behavior, but slow sorption kinetics and axial diffusion degraded the separation performance for membranes thinner than 3×10 4 μm and residence times less than 3×10 5 s.

Purification of hepatocyte growth factor using polyvinyldiene fluoride-based immobilized metal affinity membranes: equilibrium adsorption study

Polyvinyldiene fluoride (PVDF)-based affinity membranes with immobilized copper ions were developed in this study. The resulting membranes were tested for their adsorption properties using a model protein, lysozyme, in batch mode. First, different lengths of diamine were utilized as spacer arms to immobilize the metal ions onto the membranes. It was found that the application of 1,8-diaminooctane as the spacer arm led to the highest adsorption capacity. Moreover, the effects of pH and salt concentration were investigated to distinguish the proportion of specific and nonspecific interactions. A big fraction of lysozyme adsorption capacity for the immobilized metal affinity membranes was considered to come from nonspecific electrostatic interactions, which could be reduced by increasing salt concentration. Lastly, the purification of hepatocyte growth factor (HGF) from insect cell supernatant was performed using the immobilized metal affinity membranes in batch mode. HGF was found in the elution condition using EDTA, indicating the successful purification of HGF.

Protein separation using plate-and-frame modules with ion-exchange membranes

Abstract The separation performance using plate-and-frame modules with ion-exchange membranes was investigated in this study. The proteins employed were lysozyme, BSA and γ-globulin. In the batch adsorption, the adsorption behaviors of lysozyme onto cation-exchange membranes and BSA onto anion-exchange membranes, both showing the heterogeneous adsorption, could be described more appropriately with the Suen model than the Langmuir model. In the flow process using a single plate-and-frame module, operating conditions greatly influenced the separation performance. At the elution stage, a better performance was achieved by using the cross-flow mode. Other operating conditions such as the number of membranes in a stack, the pH value at adsorption, and the mixed mode in the membrane stack also affected the separation performance. As to the two-module experiments, two different operation designs were tested: connection in tandem and connection in parallel. The tandem design resulted in a better protein recovery than the parallel design when only one kind of ion-exchange membrane was adopted. Regarding the mixed-mode effect in the tandem design, when the ion-exchange membranes suitable for the target protein were placed in the second module, the target protein could gain a longer time for adsorption and, hence, a better recovery was accomplished.

Superhydrophobic waxy-dendron-grafted polymer films viananostructure manipulation

To imitate the superhydrophobicity of salient epicuticular wax on lotus leaves (hereafter “Lotus effect”), waxy dendrons were synthesized and subsequently grafted on amine-containing polystyrenes. To achieve a low surface energy and a specific surface morphology, the waxy dendron design is composed of two parts—the focal part possessing plenty of hydrogen bonding sites, and the peripheral part rich in van der Waals forces. The enhanced van der Waals force accompanied with increasing generation of dendrons helps induce self-assembly and phase separation in the preparation process of the polymer films. By different coating processes, three different films (thin film, honeycomb-like film, and three-dimensional rod-co-valley-like film) were obtained with contact angles of 95°, 130°, and 165°, respectively. The three-dimensional rod-co-valley film samples were able to imitate the superhydrophobic property (i.e. Lotus effect), as well as utilize the built-in strong hydrogen bonds to adhere water droplets on surfaces or substrates.