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Modified Batch Isotherm Determination Method for Mechanistic Model Calibration (ACS Spring Meeting 2018)

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BIOT Talk 102 given by Tobias Hahn (GoSilico) on 2018-03-19:

The fundamental assumption of in-silico scale-up and scale-down of chromatography is that only the fluid dynamics outside the pore system change. Once a molecule enters the pore system, diffusion, adsorption, and desorption are following the same mechanism from a 96-well filter plate up to a production scale column.
To obtain adsorption isotherm parameters for column modeling, batch measurements are typically corrected using an “equivalent column volume” factor. In a per-well capacity study of filter plates prepared with a ResiQuot device, considerable well-to-well differences could be found and, most importantly, deviations from the expected equivalent column volume result in wrong predictions of column experiments.
To solve this, we present a modified computational method for fitting batch isotherms to mechanistic model equations that relies only on the applied and measured supernatant concentrations. An assumption on the resin amount in the well is not needed anymore. To this, the isotherm equation is reformulated to include the liquid-to-solid ratio (L/S) as model parameter. Using this method, the average L/S in a 96-well plate filled with SP Sepharose FF could be determined from a single isotherm at constant ionic strength. The resulting binding capacity coincides with the average per well measurements, and the isotherm parameters could be used to predict a break-through curve on a 16ml lab-scale column.

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Modified Batch Isotherm Determination Method for Mechanistic Model Calibration (ACS Spring Meeting 2018)

  1. 1. STOP EXPERIMENTING. GO SILICO. Modified Batch Isotherm Determination Method for Mechanistic Model Calibration
  2. 2. Substitute experiments with simulations Tobias Hahn | Modified Batch Isotherm Determination Method | BIOT 102 In silico process developmentExperimental process development 2 Source: GE Healthcare Life Sciences https://www.gelifesciences.com/en/au/shop/ chromatography/chromatography-systems/ aktaprocess-system-p-06167
  3. 3. Applications 3Tobias Hahn | Modified Batch Isotherm Determination Method | BIOT 102 Facility Fit Process characterization Sequence of Unit Operations* Whole Process Optimization Scale-Up/ Scale-Down* Early Stage Late Stage/ Production Process Control Strategy Root Cause Analysis* Design Space Definition Optimal Experimental Design* * Case Studies on gosilico.com (BIOT 7) (BIOT 8) (BIOT 1) (BIOT 4)
  4. 4. Chromatography modeling • The chromatogram results form fluid dynamic and thermodynamic effects • Different models exist, that include different levels of detail 4Tobias Hahn | Modified Batch Isotherm Determination Method | BIOT 102
  5. 5. Scalability of models Tobias Hahn | Modified Batch Isotherm Determination Method | BIOT 102 Exchangeability of Isotherm Parameters*Lab scale Äkta model calibration Small scale Äkta model calibration Small scale LHS model calibration 5 * Case Study on gosilico.com
  6. 6. Batch Adsorption Isotherms and Kinetics 6Tobias Hahn | Modified Batch Isotherm Determination Method | BIOT 102
  7. 7. Column Adsorption and Kinetics 7Tobias Hahn | Modified Batch Isotherm Determination Method | BIOT 102
  8. 8. Steric Mass Action Model 8 • Steric Mass Action (Ion Exchange Chromatography) Tobias Hahn | Modified Batch Isotherm Determination Method | BIOT 102 Ionic capacity Characteristic charge Steric shielding (+ repulsion)
  9. 9. Usual workflow 9 1. Apply stock solutions with concentration c0 2. Shake 3. Centrifuge and measure supernatant concentration cp 4. Make assumptions on phase ratio 5. Calculate adsorbed concentration q 6. Estimate adsorption isotherm parameters from q/cp data 7. Obtain bad column predictions Tobias Hahn | Modified Batch Isotherm Determination Method | BIOT 102
  10. 10. Quantification based on ionic capacity Biotechnology Progress Volume 32(3)/2016 DOI: 10.1002/btpr.2228 Tobias Hahn | Modified Batch Isotherm Determination Method | BIOT 102 10
  11. 11. Poros 50HS SP Sepharose FF Distribution of equivalent column volume Biotechnology Progress Volume 32(3)/2016 DOI: 10.1002/btpr.2228 Tobias Hahn | Modified Batch Isotherm Determination Method | BIOT 102 11
  12. 12. Correction of each measurement 12Tobias Hahn | Modified Batch Isotherm Determination Method | BIOT 102 Biotechnology Progress Volume 32(3)/2016 DOI: 10.1002/btpr.2228
  13. 13. New approach with less assumptions 13 Comparing simulated and measured q(ε) with potentially wrong ε is error prone. 1. Substitute curve fitting objective by a function of c0 using the unknown ε 2. For IEX, account for the freed counter-ions during adsorption 3. Let isotherm develop from sample application (c0,0) towards equilibrium (cp,q) Tobias Hahn | Modified Batch Isotherm Determination Method | BIOT 102
  14. 14. Estimation result 14Tobias Hahn | Modified Batch Isotherm Determination Method | BIOT 102 Method qmax Uncorrected 900 g/L Histidine 1042 g/L Estimation 1008 g/L Measurement 1020 g/L
  15. 15. Principle 15 • 3 unknown isotherm parameter  3 features in the isotherm (c0,q) • Slope • Saturation • Curvature • 1 unknown material parameter (ε)  Determined from (c0,cp) Tobias Hahn | Modified Batch Isotherm Determination Method | BIOT 102
  16. 16. Summary 16 • Batch isotherm parameters can be used for column prediction • Adsorber amount in each well can be measured with histidine • Rigorous estimation had best results • Well porosity can be uniquely determined • A single salt condition was sufficient Tobias Hahn | Modified Batch Isotherm Determination Method | BIOT 102
  17. 17. STOP EXPERIMENTING. GO SILICO.

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