Evaluation of Three Media for Capture of Recombinant Human Serum Albumin from Pichia pastoris and Scale-up Using Capto MMC

Extracted from Multimodal Chromatography (PDF), GE Healthcare, 2013

Three media (Capto MMC, SP Sepharose Fast Flow, and SP Sepharose XL) were evaluated for their

ability to capture recombinant human serum albumin (rHSA, pI 5.5) from cell culture supernatant (CCS) of Pichia pastoris. The CCS was clarified by centrifugation and used directly as a feed material (conductivity 15 mS/cm) to measure dynamic binding capacities of the three media.

The calculated results (Table 4.16) show that Capto MMC gives a productivity of 19 kg/m3/h, which is approximately 3 times higher than that obtained with SP Sepharose Fast Flow and SP Sepharose XL when the diluted feed is used and more than 13 times higher than when the undiluted feed is used. The latter result is expected because the SP ligand is not salt tolerant. This example shows how high DBC at high conductivity combined with high flow velocities can improve the overall productivity of a capture step.

Scale-up with Capto MMC is straightforward (Fig 4.48).

Table 4.16. Productivity calculations for purification of rHSA from P. pastoris CCS. Flow velocities and residence times are based on the restrictions of the respective media in a large-scale column at 20 cm bed height. In all cases, 93% recovery and 70% loading safety factor were used1

  Dilution factor Capacity (g/L) Residence time (min) Max flow velocity (cm/h) Productivity (kg/m3/h) Productivity (kg/24 h)1
Capto MMC 15 mS/cm no dil 44 2 600 19 102
SP Sepharose XL 3 mS/cm 6.4 195 6 200 6.4 35
SP Sepharose XL 15 mS/cm no dil 0 6 200 0 0
SP Sepharose Fast Flow 3 mS/cm 6.4 135 6 200 6.0 33
SP Sepharose Fast Flow 15 mS/cm no dil 6 6 200 1.4 7.4

1 Assuming column dimensions of 120 cm diameter, 20 cm bed height (CV ≈ 225 L).

Straightforward scale-up from A) Tricorn 5/100 to B) AxiChrom 50 (100 times). In both cases the purification factor was 4 and the recovery was 93%.

Fig 4.48. Straightforward scale-up from A) Tricorn 5/100 to B) AxiChrom 50 (100 times). In both cases the purification factor was 4 and the recovery was 93%.

For more information on this example, see data file 11-0035-45, “Capto MMC.”

Materials

     
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