Thursday, March 15, 2012

Moving Past the Bottleneck

By Dr. Scott Rudge

Is there a bottleneck in Downstream Processing? The membrane chromatography vendors certainly want you to think so.

The problem is in the efficiency of chromatographic purification.  Without a doubt, chromatography is slow and inefficient. A typical protein loading for commercial scale chromatography is 25 to 40 g/L, and a typical cycle is on the order of 8 hours.  So the productivity of a chromatography column is 3 to 5 g/L/hr.  Compare this to an aggressive microbial fermentation, which produces 10 g/L in a 40 hour fermentation (0.25 g/L/hr) or a very aggressive cell culture which produces 10 g/L of antibody in seven days (0.06 g/L/hr). Clearly, even with the inefficiency of chromatography, there is plenty of volumetric productivity to keep up with modern cell culture and fermentation.

As was pointed out in a previous blog, there is no total capacity difference between typcal chromatography resins and derivatized membranes, and the dynamic binding capacity, where membrane chromatography should be superior, is also not different.So membrane chromatography does not appear to be the answer.

One technology that could intensify the performance of chromatography is “simulated moving bed” chromatography. With simulated moving beds (or SMB), the non-productive volumes of the chromatography column are put into use. This is done by segmenting the column, or making a series of much smaller columns, each of which can be operated differently at any given time.  For example, a section of the column near the classic “inlet” would be regenerated after the product has passed through it.

A section of the column downstream of the product front would be equilibrated just before the product front entered it.

In its simplest form, the SMB column is thought of in four sections, one for feed, one for product, one for regeneration, and one for raffinate, as shown in the figure below:

(from Imamoglu, S., Advances in Biochemical Engineering/Biotechnology, Vol. 76, Springer-Verlag, Berlin, p 212 (2002).)

The flow of mobile phase moves countercurrent to the direction of switching of the columns, and the velocity of switching the columns is in between the velocity of the product and the next fastest or slowest contaminant.  In the configuration shown above the raffinate moves more quickly than the product (extract), and as the solid moves counterclockwise, the extract moves backwards to the elution zone.  Meanwhile, the fast moving raffinate is allowed to exit the loop to waste.  Column segments in Zone IV are regenerated and re-equilibrated to a condition where the extract is bound but the raffinate continues to travel down column. SMB can increase the productivity of chromatography resin by a large factor.  In the simplistic diagrammatic example, the productivity could be increased by a factor of 4.  Depending on the length of the zone required for separation, the increase can be much higher.

SMB has been used for the production of amino acids, enantiomers and many other small molecules. More recently, it has been used for purification of proteins such as albumin, antibodies and some artificial demonstration mixtures such as myoglobin/lysozyme.  Innovations such as the application of gradients to SMB have been developed.  This technology has the potential to reduce cycle times and increase efficiency by smarter use of existing resources.