THE PURIFICATION OF A THERAPEUTIC POLYPEPTIDE
The purification of polypeptides is generally challenging because of the presence of many impurities that are very similar
to the target molecule. An average polypeptide consists of 20–40 amino acids and many of the impurities have a variation in
just one amino acid compared to the target molecule. For such purification problems, reversed-phase (RP) chromatography has
emerged as the major tool and it is often applied two or three times sequentially with different eluents (different buffer
species, different organic solvents).
For this work, a single RP-chromatography step of a polypeptide has been investigated. The current, optimized purification
process which is operated with conventional batch chromatography, has been taken as a benchmark. The aim of the present work
was to use the same eluent and stationary phase used in the batch setup and apply it to a continuous chromatographic process,
i.e., the MCSGP process, to show the benefits of the latter process in terms of yield, productivity, and solvent consumption.
The specifications in terms of the overall purity and the largest single impurity were equal for both processes.
For the purification of the polypeptide, a benchtop MCSGP unit was used with three columns in C18 stationary phase. Each column
was 0.75 cm i.d. x 5 cm to minimize the amount of crude material needed. The operating conditions for the MCSGP-process, including
flow rates, gradients, and switch times, were calculated following an explicit procedure that uses mainly information from
existing batch elution chromatograms.1 After the operating conditions were set, the unit was started and samples taken at regular time intervals. As with every
continuous process, the MCSGP process undergoes a transient phase until it reaches a steady state. This can be observed by
the fact that the outlet concentrations, averaged over one cycle, are constant over time. Using the steady state values regarding
productivity, yield, and solvent consumption, the comparison with the batch chromatographic process can be carried out. An
overview of the experimental results is shown in Figure 3.
All experimental results of the batch and the MCSGP processes are compared in terms of yield and productivity, under the constraint
that the same purity specifications are met. The productivity values are given as mass of purified target product per time
and per column volume. The squares in Figure 3 show the experimental results of the MCSGP process. Some variations of the
default operating points have been investigated, such as different load volumes, different switch times, etc., so the performance
in terms of yield and productivity is different. The circles with the error bars in Figure 3 indicate the performance of the
current batch purification process, where the left square shows the performance of a single batch run and the right circle
shows the performance of several batches including one recycle run with impure side fractions. Consequently, the single batch
run has a lower yield compared to the batch-plus-recycle run, but it has a slightly higher productivity. This is because the
same material has to be handled twice to run the recycle chromatography, which reduces productivity.
Figure 3 shows that for the purification problem presented, the productivity of the best MCSGP operating point is about 25
times higher than for the current batch purification process. In terms of yield, all operating points of the MCSGP process
showed higher values than the batch-plus-recycle purification scheme. For the best operating points, solvent consumption could
be reduced by 60%.
To exploit such a performance improvement in the chromatography step in production, various scenarios may be possible. In
the most likely scenario, the same amount of material should be produced in the same amount of time. With an increase in productivity
of 25 times using the MCSGP process, the column volume could be reduced 25 times, e.g., instead of 50-L column in batch chromatography,
the MCSGP process would need only three columns, each with a volume of 0.7-L. This would significantly reduce the footprint
of the chromatography unit and the surrounding tanks. Together with higher yields, the cost savings this setup would offer
are very promising and indicate payback times for the MCSGP unit of less than one year.