A z² laterally-fed membrane chromatography device for fast high-resolution purification of biopharmaceuticals

Highlights

• Improved membrane chromatography device for efficient separations.

• Novel flow arrangement for scalable uniform flow.

• Comparable protein separation to a column at 40 times the flow rate.

• Higher resolution than column at comparable flow rate.

• Increase in compactness and scalability of membrane module.

• High-resolution purification of biopharmaceuticals such as monoclonal antibodies.

Abstract

We examine the use of a z² flow distribution and collection feature for enhancing the separation efficiency of a laterally-fed membrane chromatography (or LFMC) device. The new device is designated z²LFMC. Two devices were fabricated using two different ion-exchange membranes: strong anion exchange (Q) with 0.8-micron pore size, and strong cation exchange (S) with pore size in the 3–5 µm range. The number of theoretical plates per unit membrane bed height in these z²LFMC devices were higher than those reported for other membrane chromatography devices housing similar membranes, including the older versions of LFMC devices. The enhancement in separation efficiency is explained based on computational fluid dynamic (CFD) simulations. Model protein separation experiments showed that the z²LFMC device gave similar resolution as an equivalent resin-packed column, at 40-time greater flow rate than that used with the column. At a comparable flow rate, the resolution obtained with the z²LFMC device was significantly higher than that obtained with the equivalent resin-packed column. Therefore, the z²LFMC device combines high-speed with high-resolution. We demonstrate the suitability of the z²LFMC devices for carrying out fast and efficient biopharmaceutical separations through two case studies, i.e. the fractionation of monoclonal antibody charge variants, and monoclonal antibody aggregates separation.

Introduction

Membrane chromatography (MC) is a rapid and scalable alternative to conventional resin based column chromatography [1], [2], [3], [4], [5], [6]. As solute binding takes place on the walls of flow-through pores within the membrane, the predominant mode of solute mass transport in MC is convection [1], [2], [3], [4], [5], [6]. The large frontal area in MC devices allow these to be operated at low pressures. Therefore, MC is more than an order of magnitude faster than resin based column chromatography [3]. The predominance of convection also makes separation by MC relatively less sensitive to the flow rate [3,4,7,[8], [9], [10]. Another significant advantage of MC is its suitability for single-use applications [11], [12], [13], [14], which eliminates the need for time-consuming and labour-intensive steps such as cleaning and validation [4,5,7,10,15]. Unlike a resin-packed column, whose performance is critically dependant on packing skills [16], membrane chromatography devices of consistent quality can easily be mass produced [15,17,18]. However, one of the major limiting factors with conventional MC devices such as the stacked disc and the radial flow devices is poor resolution [19], [20], [21]. For instance, it has been shown that a radial flow MC device cannot be used for carrying out high-resolution, multicomponent bind-and-elute separations [20]. This limitation could be attributed to the non-uniform flow within the device [19], [20], [21]. Resolution in MC could potentially be enhanced through the use of improved devices such as the laterally-fed membrane chromatography (or LFMC) device [19], [20], [21], [22], [23], [24], [25], [26]. A head-to-head comparison study between an LFMC device and its equivalent commercial radial flow MC device showed that while the latter was totally unsuitable for carrying out high-resolution bind-and-elute separation, multi-component mixtures could be very efficiently fractionated using the equivalent LFMC device [20]. Another study showed that even in the flow-through mode, an LFMC device outperformed its equivalent radial flow MC device [21].

An LFMC device relies on a tapered inlet for flow distribution, and a similarly tapered outlet for flow collection. While this worked well in the devices tested in our earlier studies, we recognize that a tapered inlet and outlet combination could potentially become a limiting factor while scaling up LFMC devices. Also, the separation performance of the LFMC device could potentially be enhanced further through design improvements. In a recent paper [27], we have described a novel z² flow distribution and collection feature for improving the efficiency, compactness and scalability of a resin-based cuboid packed-bed chromatography device. The overall flow path within the z² cuboid device could be visualized as a combination of two z-patterns. This z² feature eliminated the need for a tapered inlet and outlet in the cuboid packed-bed device, led to significant improvement in uniformity of flow, and resulted in improvement in separation efficiency [27]. In our current study, we examine if the separation efficiency of an LFMC device could be enhanced by incorporating the z² flow distribution and collection feature. We also describe a method for fabricating such a device.

The new LFMC device discussed in this paper is designated z²LFMC. Flow distribution and collection was achieved using primary and secondary flow channels as shown in Fig. 1A. The flow of fluid within the z²LFMC device could be classified at three levels of hierarchy, i.e. primary flow in the primary channels, secondary flow in secondary channels and tertiary (or normal) flow within the membrane stack. This three-level hierarchy was sustained by maintaining the hydraulic resistance in the membrane stack significantly higher than that in the secondary channels, which in turn was higher than that in the primary channels [27]. The arrangement of the flow channels in the device ensured that all hypothetical flow paths were of identical length. Also, the net hydraulic resistance along any hypothetical pathway was identical [27]. Hence, the residence time along any of these pathways could be expected to be the same. The use of straight flow channels in the z²LFMC device would also minimize back mixing. Therefore, the separation efficiency of an LFMC device could potentially be enhanced by adding the z² flow distribution and collection feature. Also, the uniformity in path length in a z²LFMC device would be independent of the dimensions of the device. Therefore, such a device is expected to be highly scalable. Due to the asymmetric manner in which liquid is introduced to and removed from a z²LFMC device, a tracer solute front would tilt with respect to the directions of fluid flow along the x and z axes. Fig. 1B and C show the shape of the expected solute fronts along the x-y and y-z planes respectively, within a membrane stack housed in a z²LFMC device. This combination of slants is likely to help collimate the solute front at the outlet, thereby contributing towards the narrowing of the solute residence time distribution within the device [27]. z²LFMC devices having two different bed volumes (i.e. 5 and 15 mL) were fabricated using two different types of membranes. The 5 mL device housed a stack of strong anion exchange (Q) membrane with 0.8-micron pore size, while the 15 mL device housed a stack of strong cation exchange (S) membrane having pore size in the 3–5 µm range. These devices were first characterized in terms of their number of theoretical plates. The performance of a z²LFMC device was compared with an equivalent LFMC device having tapered inlet and outlet using computational fluid dynamic (CFD) simulations. The protein separation performance of the 5 mL Q z²LFMC device was compared with a 5 mL QFF resin based column, while the performance of the 15 mL S z²LFMC device was compared with a 15 mL Capto S ImpAct resin-packed column. The 15 mL S z²LFMC device was then used for a couple of biopharmaceutical purification case studies, i.e. the fractionation of monoclonal antibody charge variants, and the separation of monoclonal antibody aggregates. The results obtained are discussed.