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Finding the best separation for enantiomeric mixtures
Finding the best separation for enantiomeric mixtures
Finding the best separation for enantiomeric mixtures
Finding the best separation for enantiomeric mixtures
Finding the best separation for enantiomeric mixtures
Finding the best separation for enantiomeric mixtures
Finding the best separation for enantiomeric mixtures
Finding the best separation for enantiomeric mixtures
Finding the best separation for enantiomeric mixtures
Finding the best separation for enantiomeric mixtures
Finding the best separation for enantiomeric mixtures
Finding the best separation for enantiomeric mixtures
Finding the best separation for enantiomeric mixtures
Finding the best separation for enantiomeric mixtures
Finding the best separation for enantiomeric mixtures
Finding the best separation for enantiomeric mixtures
Finding the best separation for enantiomeric mixtures
Finding the best separation for enantiomeric mixtures
Finding the best separation for enantiomeric mixtures
Finding the best separation for enantiomeric mixtures
Finding the best separation for enantiomeric mixtures
Finding the best separation for enantiomeric mixtures
Finding the best separation for enantiomeric mixtures
Finding the best separation for enantiomeric mixtures
Finding the best separation for enantiomeric mixtures
Finding the best separation for enantiomeric mixtures
Finding the best separation for enantiomeric mixtures
Finding the best separation for enantiomeric mixtures
Finding the best separation for enantiomeric mixtures
Finding the best separation for enantiomeric mixtures
Finding the best separation for enantiomeric mixtures
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Finding the best separation for enantiomeric mixtures

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Finding the best separation for enantiomeric mixtures

Finding the best separation for enantiomeric mixtures

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  • Speak the line, “Can we achieve this with a minimal number of columns and efficient screening strategies?”
  • Speak, “Analytical, but also preparative considerations.”
  • Speak, « Looking for a given elution order »
  • Speak that these are Polysacchasride-derived CSPs Also speak: « These will irreversibly destroy coated CSPs »
  • Speak « These will irreversibly destroy coated CSPs »
  • Speak, “An alternative possibility would be using polar organic mode to be water compatible.”
  • Different substituents New enantiorecognition
  • Transcript

    • 1. Finding the best separation for enantiomeric mixtures Chiral Technologies Europe Illkirch Cedex – France Chiral Technologies, Inc. West Chester, PA
    • 2. Analytical Method Development Goals <ul><li>Resolution of the enantiomeric pair </li></ul><ul><li>Resolution from impurities </li></ul><ul><li>Short analysis time </li></ul><ul><li>Suitable elution order </li></ul><ul><li>Compatibility of the sample media with mobile phase and column </li></ul><ul><li>Low LOD/LOQ </li></ul><ul><li>Reproducibility and robustness of the method </li></ul><ul><li>Stability of the sample under analytical conditions </li></ul>
    • 3. Method Development Overview for Polysaccharide-derived CSPs Racemic mixture HPLC SFC Organic mobile phases Water compatible mobile phases CHOICE OF SEPARATION MODE CHOICE OF TECHNOLOGY
    • 4. Choice of LC and SFC Technique <ul><li>Recognition behavior </li></ul><ul><li>Timing and productivity </li></ul><ul><li>Solubility considerations </li></ul><ul><li>Stability issues </li></ul><ul><li>Amount of mixture to be separated </li></ul><ul><li>Solvent volume to be evaporated </li></ul><ul><li>Equipment availability </li></ul>09/01/2007 00:05:36 09/01/2007 00:52:44 09/01/2007 00:29:10 -205.0 256.0 717.0 1178.0 1639.0 2100.0 Cyclique Cyclique signal UV (PIC04002:ao_F_mes_UV_DETECTOR) signal UV (PIC04002:ao_F_mes_UV_DETECTOR) (09/01/2007 00:05:36) -17.1 mv (09/01/2007 00:05:36) -17.1 mv -42.4 mv (0 jours, 00:47:09) (09/01/2007 00:52:44) -59.5 mv (09/01/2007 00:52:44) -59.5 mv -42.4 mv (0 jours, 00:47:09)
    • 5. Choice of LC and SFC Technique <ul><li>SFC Advantages of Small Scale Separations: </li></ul><ul><li>Resolution of the enantiomeric pair </li></ul><ul><li>Less solvent to be evaporated </li></ul><ul><li>Single solvent </li></ul><ul><li>Short retention times </li></ul><ul><li>Enhanced solubility in certain cases </li></ul><ul><li>Less perturbance when injecting in different solvent </li></ul>
    • 6. HPLC method development in organic mobile phases
    • 7. Polysaccharide-derived Columns <ul><li>Amylose-based </li></ul><ul><li>CHIRALPAK AD-H (Coated) </li></ul><ul><li>CHIRALPAK AS-H (Coated) </li></ul><ul><li>CHIRALPAK AZ-H (Coated) </li></ul><ul><li>CHIRALPAK AY-H (Coated) </li></ul><ul><li>CHIRALPAK IA (Immobilised) </li></ul>Cellulose-based CHIRALCEL OD-H (Coated) CHIRALCEL OJ-H (Coated) CHIRALCEL OZ-H (Coated) CHIRALPAK IB (Immobilised) CHIRALPAK IC (Immobilised)
    • 8. Primary Screening <ul><li>Immobilised </li></ul><ul><li>CHIRALPAK IA </li></ul><ul><li>CHIRALPAK IB </li></ul><ul><li>CHIRALPAK IC </li></ul><ul><li>Coated </li></ul><ul><li>CHIRALPAK AD-H </li></ul><ul><li>CHIRALCEL OD-H </li></ul><ul><li>CHIRALPAK AS-H </li></ul><ul><li>CHIRALCEL OJ-H </li></ul>
    • 9. Primary Screening
    • 10. Primary Screening <ul><li>Immobilised </li></ul><ul><li>Alkane / EtOH </li></ul><ul><li>Alkane / 2-PrOH </li></ul><ul><li>MtBE mixtures </li></ul><ul><li>Dichloromethane (or THF) mixtures </li></ul><ul><li>Coated </li></ul><ul><li>Alkane / EtOH </li></ul><ul><li>Alkane / 2-PrOH </li></ul><ul><li>EtOH / MeOH </li></ul><ul><li>Acetonitrile </li></ul>
    • 11. Basic Molecule – Analytical Scale IA IB IC Hept/EtOH/DEA Hept/IPA/DEA Hept/THF/DEA Hept/DCM/DEA Separation Retention time > 30 min Separation Retention time > 30 min 70/30/0.1 70/30/0.1 90/10/0.1 80/20/0.1 Minutes 0 2 4 6 8 10 12 14 16 18 20 22 24 mAU -50 0 50 100 150 200 250 300 350 400 450 mAU -50 0 50 100 150 200 250 300 350 400 450 1: 290 nm, 4 nm CE070128 - IA - Heptane-EtOH - 70-30-Rep2 Retention Time Minutes 0 2 4 6 8 10 12 14 16 18 20 22 24 mAU 0 50 100 150 200 250 300 mAU 0 50 100 150 200 250 300 1: 290 nm, 4 nm CE070128 - IA - Heptane-IPA - 70-30-Rep2 Retention Time Minutes 0 2 4 6 8 10 12 14 16 18 20 22 24 mAU -2,5 0,0 2,5 5,0 7,5 10,0 12,5 15,0 17,5 20,0 22,5 25,0 mAU -2,5 0,0 2,5 5,0 7,5 10,0 12,5 15,0 17,5 20,0 22,5 25,0 1: 290 nm, 4 nm CE07128 - IA - Hept-THF 90-10-Rep2 Retention Time Minutes 0 2 4 6 8 10 12 14 16 18 20 22 24 mAU -5 0 5 10 15 20 25 30 mAU -5 0 5 10 15 20 25 30 1: 290 nm, 4 nm CE07128 - IA - Hept-DCM 80-20-Rep2 Retention Time Minutes 0 2 4 6 8 10 12 14 16 18 20 22 24 mAU -50 0 50 100 150 200 250 300 350 400 450 mAU -50 0 50 100 150 200 250 300 350 400 450 1: 290 nm, 4 nm CE070128 - IB - Heptane-EtOH - 70-30-Rep2 Retention Time Minutes 0 2 4 6 8 10 12 14 16 18 20 22 24 mAU -25 0 25 50 75 100 125 150 175 200 225 250 275 mAU -25 0 25 50 75 100 125 150 175 200 225 250 275 1: 290 nm, 4 nm CE070128 - IB - Heptane-IPA - 70-30-Rep2 Retention Time Minutes 0 2 4 6 8 10 12 14 16 18 20 22 24 mAU -50 0 50 100 150 200 250 300 350 400 450 mAU -50 0 50 100 150 200 250 300 350 400 450 1: 290 nm, 4 nm CE070128 - IC - Heptane-EtOH - 70-30-Rep2 Retention Time Minutes 0 2 4 6 8 10 12 14 16 18 20 22 24 mAU -25 0 25 50 75 100 125 150 175 200 225 250 mAU -25 0 25 50 75 100 125 150 175 200 225 250 1: 290 nm, 4 nm CE070128 - IC - Heptane-IPA - 70-30-Rep2 Retention Time Minutes 0 2 4 6 8 10 12 14 16 18 20 22 24 mAU -2,5 0,0 2,5 5,0 7,5 10,0 12,5 15,0 17,5 20,0 22,5 25,0 mAU -2,5 0,0 2,5 5,0 7,5 10,0 12,5 15,0 17,5 20,0 22,5 25,0 1: 290 nm, 4 nm CE07128 - IC - Hept-THF 90-10-Rep2 Retention Time Minutes 0 2 4 6 8 10 12 14 16 18 20 22 24 mAU -10 0 10 20 30 40 50 mAU -10 0 10 20 30 40 50 1: 290 nm, 4 nm CE07128 - IC - Hept-DCM 80-20-Rep2 Retention Time
    • 12. Basic Molecule – Analytical Scale CHIRALPAK IB n -heptane/2-PrOH/DEA 70/30/0.1 12 min 7 min
    • 13. Complementary Properties of CSPs <ul><li>CHIRALPAK IA </li></ul><ul><li>CHIRALPAK IB </li></ul><ul><li>CHIRALPAK IC </li></ul>(D) (L) min 0 2.5 5 7.5 10 12.5 15 17.5 (D) (L) FMOC-D,L-Leucine n -hexane/2-PrOH/TFA 90/10/0.1 (D) min 0 2.5 5 7.5 10 12.5 15 17.5 (L) min 0 2.5 5 7.5 10 12.5 15 17.5
    • 14. Complementary Properties of CSPs <ul><li>Advantages </li></ul><ul><li>Specific resolutions </li></ul><ul><li>Changes in enantiomeric elution order </li></ul><ul><li>Different elution of impurities </li></ul>
    • 15. SFC method development in organic mobile phases
    • 16. Compatible Co-Solvents in SFC <ul><li>Coated CSPs </li></ul><ul><li>Ethanol </li></ul><ul><li>Methanol </li></ul><ul><li>2-Propanol </li></ul><ul><li>Acetonitrile </li></ul><ul><li>Other alcohols </li></ul><ul><li>Immobilised CSPs </li></ul><ul><li>Ethanol </li></ul><ul><li>Methanol </li></ul><ul><li>2-Propanol </li></ul><ul><li>Acetonitrile </li></ul><ul><li>Other alcohols </li></ul>
    • 17. Compatible Co-Solvents in SFC <ul><li>Extended Solvent Range </li></ul><ul><li>MtBE </li></ul><ul><li>Ethyl acetate </li></ul><ul><li>THF </li></ul><ul><li>Dichloromethane </li></ul><ul><li>Chloroform </li></ul><ul><li>1,4-dioxane </li></ul><ul><li>Acetone </li></ul><ul><li>DMSO or DMF (as injection solvents) </li></ul>
    • 18. CSPs in SFC: Analytical Scale CO 2 / MeOH 90/10 T=25°C Flow rate: 3.0ml/min. CHIRALPAK IA CO 2 / THF (+1%DEA) 70/30 T=30°C Flow rate: 3.0ml/min. CHIRALPAK IC 0 200 400 600 800 1000 1200 (sec)
    • 19. CSPs in SFC: Analytical Scale <ul><li>All the advantages of the SFC applications on polysaccharide-derived CSPs </li></ul><ul><li>Possibility of eluting and/or injecting in different solvents (DCM, THF, …) when using the immobilised CSPs </li></ul>
    • 20. HPLC method development in water compatible mobile phases
    • 21. Water Compatible Mobile Phases Aqueous Solution Organic modifier Identify the compound nature Acidic Neutral Basic HCOOH pH 2.0 40% ACN 60% MeOH H 2 O 40% ACN 60% MeOH 20mM NH 4 HCO 3 pH 9.0 40% ACN 60% MeOH
    • 22. Separations RP-Mode CHIRALPAK IA furoin MeOH/H 2 O 55:45 min 0 2 4 6 8
    • 23. Separations RP-Mode CHIRALPAK IC 2,3-dibenzoyl- DL-tartaric acid HCOOH aq. (pH2.0) ACN: 55% min 0 2 4 6 8
    • 24. Faster screening and enhanced resolution: 3-µm CSPs
    • 25. Fast Analysis: CHIRALPAK IA-3 (4.6 x 50 mm) Hexane-EtOH 80:20 (+0.1% AE) Flow rate: 5.0 ml/min DCM-MeOH 98:2 (+0.1% AE) Flow rate: 5.0 ml/min MeOH 100% Flow rate: 5.0 ml/min 40 seconds 30 seconds 20 seconds
    • 26. Looking for new separations: different chiral selectors
    • 27. Different Substituents; New Enantiorecognition CHIRALCEL OD CHIRALPAK AD CHIRALCEL OZ CHIRALPAK AZ CHIRALPAK IC CHIRALPAK AY
    • 28. Are New Selectors Needed? Metolachlor Hexane / EtOH 95:5 1ml/min, 25°C (4.6 x 250 mm) CHIRALPAK IA CHIRALPAK IB CHIRALPAK IC CHIRALPAK IA CHIRALPAK IB CHIRALPAK IC
    • 29. Are New Selectors Needed? Metolachlor Hexane / EtOH 95:5 1ml/min, 25°C (4.6 x 250 mm) CHIRALPAK AS-H CHIRALPAK AD-H CHIRALCEL OD-H CHIRALCEL OJ-H
    • 30. Are New Selectors Needed? Metolachlor Hexane / EtOH 95:5 1ml/min, 25°C (4.6 x 250 mm) CHIRALPAK AY-H
    • 31. Chiral Technologies Worldwide <ul><li>Quality </li></ul><ul><li>Expertise </li></ul><ul><li>Solutions </li></ul>www.chiraltechworldwide.com Americas: 610-594-2100 Europe: 00 33 (0)3 88 79 52 00

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