CONTINUOUS PROCESSING: The Multicolumn Countercurrent Solvent Gradient Purification Process - A continuous chromatographic process for monoclonal antibodies without using Protein A - BioPharm

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CONTINUOUS PROCESSING: The Multicolumn Countercurrent Solvent Gradient Purification Process
A continuous chromatographic process for monoclonal antibodies without using Protein A


BioPharm International


To facilitate the following explanation, the time axis of the chromatogram (Figure 1) has been reversed (i.e., it goes from right to left). The generic chromatogram can be cut into five fractions as indicated by the numbers on the time axis:
1 = strongly adsorbing impurities
2 = product contaminated by strongly adsorbing impurities
3 = product
4 = product contaminated with weakly adsorbing impurities
5 = weakly adsorbing impurities.

The aim of an ideal purification process would be to collect fraction 3, to drain fractions 1 and 5, and to recycle fractions 2 and 4. Transferring these tasks to a multicolumn chromatographic process results in a 6-column flowsheet (Figure 2), which shows the basic principle of the MCSGP process.


Figure 2. Basic principle of the MCSGP process
Figure 2 shows six columns. Each column is connected to a gradient pump. The outlet of columns 2 and 4 is recycled into columns 4 and 6, respectively. Fractions 1 to 5 as shown in the chromatogram in Figure 1 are eluted from a column with the same number (Figure 2). For example, the strongly adsorbing impurities (i.e., fraction 1 from Figure 1), are drained from column 1. In column 2, the product fraction contaminated with strongly adsorbing impurities, P+S (i.e., fraction 2) is eluted, mixed with the outlet of the gradient pump, and recycled to column 4. In column 3, the product is collected. The outlet of column 4, P+W, is recycled to column 6. In column 5, the raw feed mixture is fed to the column inlet and the weakly adsorbing impurities are eluted. In order to run this process continuously, after a certain time all columns are switched one position to the left (e.g., column 4 becomes column 3), and column 1, being void of any solutes at the end of the switch time, can be switched to the position of column 6.3–4 This column switching results in a countercurrent movement of liquid and solid flow. This countercurrent operation improves the efficiency of the process.


Figure 3. Flowsheet of the MCSGP process
To reduce equipment costs, the MCSGP process used with just three columns. This reduction from six to three columns can be achieved by performing the draining or collecting of S, P, and W and the recycling of P+S and P+W, not in parallel (Figure 2), but in two sequential steps. This results in two separate flowsheets, which are applied to the three columns in an alternating fashion (Figure 3).

After the columns have been switched one position to the left (where column 1 is switched to the position of column 3), the columns are operated in the countercurrent mode (i.e., recycling of P+S and P+W into the subsequent column takes place). After a certain time, t CC , the columns are connected in a different manner and then operated for a time, t B , in the batch mode (i.e., while the draining of S and W and the collecting of P take place). Then the columns are switched again and operate in the countercurrent mode. The column switching plus countercurrent and batch operation mode makes one cycle of time (t CC + t B ) and this cycle is repeated.


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