Design of RP-HPLC 1 for Insulin Production Presented by: Patricia Yee, Nicholas Irianto, Thomas Fang Overview Background Information Significance of RP-HPLC1 Design Parameters Selected Design and Specifications Questions Background Information What is HPLC? High Performance Liquid Chromatography - or - High-Pressure Liquid Chromatography How HPLC works… Reverse-Phase HPLC Insulin Production Process: RP-HPLC 1 Insulin Production Process: RP-HPLC 1 What comes out… 1) Contaminated protein, denatured insulin, enzyme and small amounts of insulin ester and WFI 2) Majority of desired insulin ester and WFI Insulin Production Process: RP-HPLC 1 Gradient Elution of mobile phase: Acetonitrile / Water / Acid Main Design Parameters Column Size(Internal diameter) Material (Stainless Steels, Glass, etc) Main Design Parameters Resin Particlesize (10μm ~75μm ) Pore size (300 Á) Material (Silica, styrene-divinylbenzene, etc.) Other Design Considerations Pump (pressure capacity) Solvent Acetonitrile Polar solvent (increase retention time) Hydrophobic solvent (decrease retention time) Possible Design Design Specifications •100% binding and 90% yield for insulin ester •60% binding and 5% yield for denatured insulin precursors •Purification results in outlet stream consisting of 340 kg insulin ester/batch Design Selections HPLC Columns •Novasep Prochrom® DAC Columns LC1600 H x D: 450 x 1600 mm HPLC Resins •PRP-3 standard Poly(styrene- divinylbenzene) (PSDVB) Pore size 300 Å Diameter 10 μm Design Justifications Stable from pH 1-13 Does not contain acidic silanol groups, which are present on silica supports. Highly cross-linked for exceptional durability and to prevent shrinking and Consistent performance 10μm particle diameter is selected for better solution and peak shape Design Details Twenty three LC 1600 HPLC column units with 0.91 hours process time would be required for the process according to SuperPro For one LC 1600 HPLC column with maximum bed volume of 900 L would require 360 000 grams of PRP-3 resins COSTS $24/g of PRP-3 resins This translates to $ 8 640 000 / LC 1600 HPLC column For 23 HPLC columns, it would cost $ 198 720 000 $28 000 for a Novasep Prochrom DAC LC 1600 column Total Cost $ 200 000 000 Questions? THANK YOU!