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Anion Exchanger

Anion Exchanger. Kristen Galea. Where Are We With in the Process?. Entering stream is concentrated with, insulin ester, and denatured proteins This separation technique separates anionic insulin ester from denatured ester and proteins Two streams leave process Impurities

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Anion Exchanger

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  1. Anion Exchanger Kristen Galea

  2. Where Are We With in the Process? • Entering stream is concentrated with, insulin ester, and denatured proteins • This separation technique separates anionic insulin ester from denatured ester and proteins • Two streams leave process • Impurities • Insulin ester for hydrolysis reaction

  3. Principles of Ion Exchange • Charged resin beads packed in a column • Proteins stick to resins by electrostatic interactions • If solution is below the isoeletric point of the protein, cation exchange will occur (protein will have net + charge) • If solution is above the isoeletric point of the protein, anion exchange will occur (protein will have net – charge)

  4. Principles of Ion Exchange • Proteins will be exchanged with the free ion around the resin beads – binding will occur • Elution of protein involves a change in pH (or addition of a salt or solvent) • Changes the charge on the protein – release of the protein

  5. Anion Exchange Resins • Insoluble polymeric beads • Manufactured by polymerization of styrene and divinylbenzene (DVB) • DVB acts as a crosslinking agent and without styrene would be water soluble • Active groups (+) are covalently attached to bead balanced by equivalent number of oppositely charged (-) free ions

  6. Anion Exchanger • 3 steps to Anion Exchange • Binding • Wash • Elution

  7. Anion Exchanger • Binding – inlet to column runs through • 100% of insulin esters bind to resin beads • Wash with NaOH/Water (high pH) several times • Removes everything except insulin ester • Elute column with NaOH/Water (pH close to pI of protein – 5.4) • Insulin ester changes from a negative charged to neutral • Released from beads – 90% yield

  8. Anion Exchange • Elution process simultaneously replenishes the negatively charged free ions around the resin (OH-) • Purified insulin ester stream then undergoes hydrolysis • Multiple columns to make a continuous process

  9. Proposed Design • Assumptions • Batch = 24 hours • Flow entering = 67.9 kg/h • Bed Capacity = 0.01 kg/L • Estimated Dimensions • Bed Volume = 300 L (selected) • Bed Diameter = 1.5 m • Bed Height = 0.17 m • Diameter larger than height of bed, to reduce pressure drop

  10. Suppliers • Column • Difficult to find – must be custom made • Amersham Biosciences • Resin • Based on what is being removed • Must have high capacity to retain proteins

  11. Suppliers • Resin • Amersham Biosciences – Source 15Q • Strong Anion Exchange resin • Good for protein separation • Problem • Binding Capacity: 4.5X10-3kg/L • Low flow rate

  12. Alternatives • Different types of elution can be used, however, more sophisticated control systems must be used to ensure appropriate amount of protein is removed • More testing required for this process.

  13. Questions?

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