Polymorph Quantitation

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Investigations in Pharmaceutical Polymorph Quantification using PXRD, ATR, FTIR, and FT-Raman at Roche Palo Alto

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Polymorph Quantitation

  1. 1. Investigations in Pharmaceutical Polymorph Quantification using PXRD, ATR‒FTIR, and FT‒Raman at Roche Palo Alto Richard E. Young Research Scientist II Analytical Research Roche Palo Alto LLC
  2. 2. Agenda  Basic Principles of Polymorphism  Polymorphism’s Importance to Pharmaceuticals  Polymorph Discovery Techniques and the Allied Analytical Tools  Case Study 1: Three Phases of Gancilclovir  Case Study 2: Two Phases of a Roche Research Compound  Conclusion 12/30/2009 Richard E. Young 2
  3. 3. What is a Chemical Polymorph? A compound with a single molecular structure that possess more than one crystal form. A polymorph form is often termed a “phase.” http://en.wikipedia.org/wiki/Bravais_lattice 12/30/2009 Richard E. Young 3
  4. 4. Differentiating Habit and Crystal Chemistry of a Compound J. K. Haleblian, “Characterization of habits and crystalline modification of solids and their pharmaceutical applications,” J. Pharm. Sci., 64(8), 1270 (1975). Chemical Compound (Solid) Habit Internal Structure Crystalline Amorphous Single Entity Molecular Adduct Polymorphs Nonstoichiometric Stoichiometric Inclusion compounds Solvates (hydrates) Channel Layer Cage (clathrate) 12/30/2009 Richard E. Young 4
  5. 5. The Crystallinity Continuum The “crystallinity continuum” is expressed by the relative amount of order of contiguous unit cells High Large single crystal Crystalline order Powdered crystals Small crystallites (more or less ordered) Semi-crystalline (short range order) Low Amorphous (lacks a distinct crystal form ) 12/30/2009 Richard E. Young 5
  6. 6. The Importance of Polymorphism in Pharmaceuticals • Pharmaceutical performance Each polymorph may have different physical properties (melting point, solubility, dissolution rate, processibility, etc.), which may affect: – Stability – Formulation – Potency – Bioavailability – Storage • Intellectual Property – Each polymorph can be patented if it shows better properties than any previously patented polymorph – A rival company may legally sell the same drug substance in a different crystal form if the new polymorph is not patented 12/30/2009 Richard E. Young 6
  7. 7. Polymorph Discovery Techniques No method can predict polymorphs. Discovery must be done by empirical techniques. • Stability studies with varying temperature, humidity, and time • Re-crystallization with different solvents • Melt re-crystallization • Anti-solvent addition • Annealing 12/30/2009 Richard E. Young 7
  8. 8. Polymorph Analytical Tools • Powder X-Ra Diffraction (PXRD or XRPD) • Thermal techniques – Differential scanning calorimetry (DSC) – Differential thermal analysis (DTA) – hermal gravimetric analysis (TGA) – Melting point – Thermal microscopy • Solid State NMR • Vibrational Spectroscopy – FTIR (ATR, microscopy, etc.) – FT-Raman – NIR 12/30/2009 Richard E. Young 8
  9. 9. The Polymorph Analytical Tools to be Examined • Powder X-Ray Diffraction • AT-FTIR • FT-Raman 12/30/2009 Richard E. Young 9
  10. 10. PXRD Bragg-Brentano Theta-Theta Configuration Mono- chromater Divergence slit Antiscatter- slit Detector slit Tube http://www.thermo.com/eThermo/CMA/PDFs/Product/productPDF_11602.pdf http://www.smcr.fisica.unam.mx/8temasutiles/articulosutiles/Bas-XRD.pdf • A crystalline solid will produce a distinctive “fingerprint” pattern of sharp peaks; amorphous materials will produce a broad “hump” • Phase characterization and identification - not optimum for structure determinations • Transmission or reflection configuration 12/30/2009 Richard E. Young 10
  11. 11. Bragg Diffraction Bragg’s Law: nλ = 2d sin θ θ is the scattering angle λ is the X-ray wavelength d is the distance between planes http://www.smcr.fisica.unam.mx/8temasutiles/articulosutiles/Bas-XRD.pdf) • A crystal modeled as a series of parallel planes with distance “d” between planes (d-spacing) • Constructive interference of reflections creates a Bragg peak 12/30/2009 Richard E. Young 11
  12. 12. Powder X-Ray Diffraction Advantages and Disadvantages • Advantages – Well established and accepted (the gold standard) – Rapid and simple sample analysis – Can readily differentiate polymorphs – Can analyze mixed polymorphs – Quantitative and qualitative • Disadvantages – Sensitive to sample preparation technique – Requires radiation license and fees to operate – X-ray hazard – Very expensive 12/30/2009 Richard E. Young 12
  13. 13. PXRD: Example of an Amorphous and Crystalline Drug Substance Mixture O O O N N O N O O F F N O O F 0% Amorphous RS-104253-146 F 30% Amorphous 50% Amorphous 70% Amorphous 100% Amorphous 3 10 20 30 40 2-Theta 12/30/2009 Richard E. Young 13
  14. 14. PXRD: Example of Polymorph Transitions by Temperature 200 °C 200 C O O Cl N S O O O O RO0130830-000 175 °C 175 C 155 °C 155 C 110 °C 110 C 30 °C 30 C 2 10 20 30 2-Theta 12/30/2009 Richard E. Young 14
  15. 15. Multi-bounce Attenuated Total Reflectance - FTIR Sample clamp Sample Multi-bounce ATR crystal IR Radiation IR Detector Mirror Mirror • Solid & liquid samples may be analyzed without preparation • Solid samples are firmly clamped against crystal to provide intimate contact reducing the distorting effect of trapped air • ATR crystals: germanium, KRS-5, zinc selenide, silicon, diamond 12/30/2009 Richard E. Young 15
  16. 16. ATR-FTIR (with diamond) Advantages and Disadvantages • Advantages – Sample holder (diamond) scratch & abrasion resistant – Rapid sample analysis – Minimal sample preparation required – Quantitative and qualitative – Can analyze mixed polymorphs • Disadvantages – Diamond absorbs in the 2300 to 1800 cm-1 region – Polymorph distinction is compound dependent – Not as well established for polymorph analysis – Expensive (diamond) 12/30/2009 Richard E. Young 16
  17. 17. Raman Scattering Scattered radiation Sample Excitation energy (laser radiation) Exited energy states Rayleigh scatter Rayleigh scattering (same wavelength as excitation energy)) Excitation energy Anti-stokes Stokes 4 Raman scatter energy states 3 Vibrational 2 (stokes & anti-stokes - different 1 wavelengths than excitation energy) 0 12/30/2009 Richard E. Young 17
  18. 18. FT-Raman Advantages and Disadvantages • Advantages – Rapid sample analysis – Minimal sample preparation required – Quantitative and qualitative – Can analyze mixed polymorphs • Disadvantages – Polymorph distinction is compound dependent – Not as well established for polymorph analysis 12/30/2009 Richard E. Young 18
  19. 19. Case Study 1: Ganciclovir 9-(1,3-dihydroxy-2-propoxymethyl) guanine • Antiviral drug for the treatment for cytomegalovirus (CMV) infections • Four known polymorphs: Phases I, II, & III and Phase I Hydrate • PXRD quantifiation method for Phases I, II, and III 12/30/2009 Richard E. Young 19
  20. 20. PXRD Patterns of Three Polymorphs of Ganciclovir Ganciclovir Phase I Phase I Ganciclovir Phase II Phase II Ganciclovir Phase III Phase III 2 10 20 30 40 2-Theta 2-Theta 12/30/2009 Richard E. Young 20
  21. 21. Equations for Calculating Ganciclovir Phase Percents using PXRD Analysis If Only Phases I & II If Only Phases I & III 100% 100% % Phase II = % Phase I = Slope 1 ( Peak I Peak II ) +1 ( Slope 2 Peak III Peak I ) +1 % Phase I = 100% - % Phase II % Phase III = 100% - % Phase I If Only Phases II & III If All Three Phases (I, II, & II) 100% 100% % Phase I = ( ) ( ) % Phase II = ( ) Peak III Peak II Peak III Slope 3 +1 Slope 1 + Slope 3 +1 Peak II Peak I Peak I % Phase III = 100% - % Phase II % Phase III = % Phase I (Peak III Peak I ) Slope 3 % Phase II = 100% - % Phase I - % Phase III 12/30/2009 Richard E. Young 21
  22. 22. Quantifation of Standards of Ganciclovir Polymorph Mixtures by PXRD (Set A) Ratios Phase I Phase II Phase III Phase I Phase II Phase III 05/95/00 -3.39 3.39 0.00 10/00/90 -1.16 0.00 1.16 45/50/05 -3.64 3.59 0.06 20/75/05 -5.49 6.23 -0.74 60/35/05 -3.78 3.40 0.38 70/05/25 -1.38 -0.48 1.86 80/10/10 -2.21 0.05 2.16 00/95/05 0.00 0.65 -0.65 95/05/00 0.06 -0.06 0.00 95/00/05 -0.45 0.00 0.45 12/30/2009 Richard E. Young 22
  23. 23. Quantifation of Standards of Ganciclovir Polymorph Mixtures by PXRD (Set B) Ratios Phase I Phase II Phase III Phase I Phase II Phase III 05/95/00 -5.75 5.75 0.00 10/00/90 0.04 0.00 1.76 45/50/05 -11.04 9.38 1.66 20/75/05 -8.21 7.35 0.87 60/35/05 -5.92 4.86 1.06 70/05/25 -5.25 -0.08 5.32 80/10/10 -2.06 0.55 1.51 00/95/05 0.00 -0.41 0.41 95/05/00 0.20 -0.20 0.00 95/00/05 0.23 0.00 -0.23 12/30/2009 Richard E. Young 23
  24. 24. ATR-FTIR Spectra of Three Polymorphs of Ganciclovir Salari, A. and R. Young, "Application of Attenuated Total Reflectance FTIR Spectroscopy to the Analysis of Mixtures of Pharmaceutical Polymorphs," International Journal of Pharmaceutics, 163, 157-166 (1998). Phase I Phase II Phase III 12/30/2009 Richard E. Young 24
  25. 25. Ganciclovir Polymorph Calibration Standards Int J of Pharm 163, 157-166 (1998) • Single Phase Standards I, II, & III • Binary Phase Standards I & II I & III II & III Three sets: 5, 10, 25, 50, 75, 90, 95 wt% each • Ternary Phase Standards I, II, & III – Six mixtures consisting of 10, 30, 60 wt% each – One mixture of 33.3 wt% each 12/30/2009 Richard E. Young 25
  26. 26. ATR-FTIR Ganciclovir Polymorph Calibrations Using Partial Least Squares Int J of Pharm 163, 157-166 (1998) Phase I r2 = 0.962 Phase II r2 = 0.964 Phase III r2 = 0.972 12/30/2009 Richard E. Young 26
  27. 27. FT-Raman Spectra of Ganciclovir Polymorphs Phase I Phase II Phase III 12/30/2009 Richard E. Young 27
  28. 28. Quantifation of Standards of Ganciclovir Phase I Polymorph by ATR & FT-Raman Int J of Pharm 163, 157-166 (1998) Actual Mean Stnd Dev Difference wt% ATR RAM ATR RAM ATR RAM ATR Raman 0.0 2.8 -1.2 4.0 2.0 -2.8 1.2 5.0 7.6 6.6 5.7 2.2 -2.6 -1.6 10.0 8.3 12.2 1.8 2.4 1.7 -2.2 25.0 21.8 26.9 3.5 1.5 3.2 -1.9 30.0 19.7 32.2 1.8 3.3 10.3 -2.2 33.3 25.2 33.5 0.8 3.8 8.1 -0.2 50.0 50.9 54.2 5.2 4.3 -0.9 -4.2 60.0 56.3 65.2 3.2 2.8 3.7 -5.2 75.0 71.6 75.9 9.2 5.2 3.4 -0.9 90.0 92.4 88.8 8.8 3.3 -2.4 1.2 95.0 95.0 93.1 1.2 3.7 0.0 1.9 100 106 99.3 3.2 3.2 -6.0 0.7 12/30/2009 Richard E. Young 28
  29. 29. Quantifation of Standards of Ganciclovir Phase II Polymorph by ATR & FT-Raman Int J of Pharm 163, 157-166 (1998) Actual Mean Stnd Dev Difference wt% ATR RAM ATR RAM ATR RAM ATR Raman 0.0 -0.9 0.0 4.3 1.6 0.9 0.0 5.0 4.0 4.5 1.3 1.1 1.0 0.5 10.0 12.5 10.0 2.8 0.7 -2.5 0.0 25.0 29.2 26.6 3.0 2.6 -4.2 -1.6 30.0 32.6 28.4 3.8 2.0 -2.6 1.6 33.3 39.8 33.0 3.0 2.7 -6.5 0.3 50.0 47.0 47.0 5.4 4.6 3.0 3.0 60.0 66.8 56.2 2.7 2.9 -6.8 3.8 75.0 73.0 76.8 8.8 2.2 2.0 -1.8 90.0 87.1 89.3 8.0 1.3 2.9 0.7 95.0 89.1 94.2 10.5 1.5 5.9 0.8 100 94.0 102 9.7 1.7 6.0 -2.0 12/30/2009 Richard E. Young 29
  30. 30. Quantifation of Standards of Ganciclovir Phase III Polymorph by ATR & FT-Raman Int J of Pharm 163, 157-166 (1998) Actual Mean Stnd Dev Difference wt% ATR RAM ATR RAM ATR RAM ATR Raman 0.0 2.9 1.30 5.6 2.3 -2.9 -1.3 5.0 4.8 6.1 1.3 2.5 0.2 -1.1 10.0 12.6 9.50 5.8 1.7 -2.6 0.5 25.0 22.2 21.8 6.9 1.9 2.8 3.2 30.0 27.3 27.4 0.6 5.3 2.7 2.6 33.3 33.9 33.5 3.2 1.5 -0.6 -0.2 50.0 48.1 47.1 6.6 3.5 1.9 2.9 60.0 62.8 60.1 7.3 2.2 -2.8 -0.1 75.0 75.4 73.4 6.4 2.9 -0.4 1.6 90.0 89.1 88.8 7.6 3.5 0.9 1.2 95.0 95.9 92.9 2.0 3.4 -0.9 2.1 100 102 107 7.7 0.4 -2.0 -7.0 12/30/2009 Richard E. Young 30
  31. 31. Ganciclovir Polymorph Validation Mixtures Int J of Pharm 163, 157-166 (1998) • One of Each Pure Phase Validation Phase I Phase II Phase III Mix ID (wt%) (wt%) (wt%) – 100 % Phase I Phase I 100 0.0 0.0 – 100% Phase II Phase II 0.0 100 0.0 – 100% Phase III Phase III 0.0 0.0 100 Mix 1 5.6 21.2 73.2 Mix 2 7.3 9.8 82.8 • Ternary Phase Mixtures Mix 3 14.2 43.1 42.7 – Ten mixtures of all three Mix 4 14.1 78.6 7.4 phases in varying Mix 5 26.8 56.0 17.2 amounts Mix 6 33.6 11.4 55.0 Mix 7 40.8 18.2 41.0 Mix 8 46.4 32.9 20.7 Mix 9 46.5 28.2 25.4 Mix 10 58.2 18.2 23.5 12/30/2009 Richard E. Young 31
  32. 32. Quantifation of Validation Mixes of Ganciclovir Phase I Polymorph by ATR & FT-Raman Int J of Pharm 163, 157-166 (1998) Validation Actual Calc wt% Difference Mix ID wt% ATR RAM ATR RAM ATR Raman Phase I 100 102 99.3 -2.0 0.7 Phase II 0.0 3.2 -2.7 -3.2 2.7 Phase III 0.0 5.6 -4.6 -5.6 4.6 Mix 1 5.6 9.0 6.9 -3.4 -1.3 Mix 2 7.3 9.0 9.7 -1.7 -2.4 Mix 3 14.2 21.4 10.2 -7.2 4.0 Mix 4 14.1 12.5 14.3 1.6 -0.2 Mix 5 26.8 23.6 27.4 3.2 -0.6 Mix 6 33.6 39.5 44.8 -5.9 -11.2 Mix 7 40.8 45.6 42.2 -4.8 -1.4 Mix 8 46.4 47.7 45.9 -1.3 0.5 Mix 9 46.5 47.2 49.9 -0.7 -3.4 Mix 10 58.2 61.5 59.7 -3.3 -1.5 12/30/2009 Richard E. Young 32
  33. 33. Quantifation of Validation Mixes of Ganciclovir Phase II Polymorph by ATR & FT-Raman Int J of Pharm 163, 157-166 (1998) Validation Actual Calc wt% Difference Mix ID wt% ATR RAM ATR RAM ATR Raman Phase I 0.0 -2.1 -0.1 2.1 0.1 Phase II 100 104 102 -4.0 -2.0 Phase III 0.0 1.2 -2.7 -1.2 2.7 Mix 1 21.2 22.0 21.7 -0.8 -0.5 Mix 2 9.8 11.1 10.0 -1.3 -0.2 Mix 3 43.1 46.0 47.8 -2.9 -4.7 Mix 4 78.6 86.8 82.4 -8.2 -3.8 Mix 5 56.0 65.9 58.4 -9.9 -2.4 Mix 6 11.4 7.2 4.9 4.2 6.5 Mix 7 18.2 22.7 17.7 -4.5 0.5 Mix 8 32.9 37.5 34.5 -4.6 -1.6 Mix 9 28.2 35.4 28.1 -7.2 0.1 Mix 10 18.2 21.1 18.1 -2.9 0.1 12/30/2009 Richard E. Young 33
  34. 34. Quantifation of Validation Mixes of Ganciclovir Phase III Polymorph by ATR & FT-Raman Int J of Pharm 163, 157-166 (1998) Validation Actual Calc wt% Difference Mix ID wt% ATR RAM ATR RAM ATR Raman Phase I 0.0 -1.1 0.8 1.1 -0.8 Phase II 0.0 -6.7 0.3 6.7 -0.3 Phase III 100 94.1 107 5.9 -7.0 Mix 1 73.2 68.5 71.4 4.7 1.8 Mix 2 82.8 80.3 80.3 2.5 2.5 Mix 3 42.7 32.2 42.0 10.5 0.7 Mix 4 7.4 0.7 3.2 6.7 4.2 Mix 5 17.2 10.2 14.1 7.0 3.1 Mix 6 55.0 54.7 50.3 0.3 4.7 Mix 7 41.0 31.6 40.0 9.4 1.0 Mix 8 20.7 14.3 19.6 6.4 1.1 Mix 9 25.4 17.0 22.0 8.4 3.4 Mix 10 23.5 17.6 22.2 5.9 1.3 12/30/2009 Richard E. Young 34
  35. 35. Summary of Case Study 1 Three Polymorphs of Ganciclovir • ATR and Raman have the capability to quantify complex mixtures of polymorphs • ATR and Raman produced quantitative results comparable to the PXRD • The Raman, in general, gave smaller differences than ATR or PXRD 12/30/2009 Richard E. Young 35
  36. 36. Case Study 2: Anhydrate and Hydrate Phases of a Roche Research Compound in Tablets Identification and quantification of two phases of a Roche research compound in the presence of excipients Research Compound & excipients (50:50) in 150-mg tablets • Anhydrate (“Phase A”) • Hydrate (“Phase B”) • Excipients (“Placebo”): Pharmatose 350M, Povidone K30, Ac-Di-Sol, Avicel PH102, magnesium stearate 12/30/2009 Richard E. Young 36
  37. 37. PXRD Patterns of Phase A, Phase B, and Placebo 12/30/2009 Richard E. Young 37
  38. 38. PXRD Patterns of Polymorph Standards (% Phase B in Phase A & Placebo) Phase A Phase B 9.0 2θ 13.1 2θ 12/30/2009 Richard E. Young 38
  39. 39. PXRD Reference Intensity Ratio (RIR) Technique for Two Polymorph Mixtures 100 % wt% B =  1 + (m × IA ÷ IB) • wt% B is the weight percent of Phase B in the sample • m is the slope of the line of the (XB / XA) vs (IB / IA) linear regression • IA is the peak height of the Phase A peak at 9.0 2θ in the sample • IB is the peak height of the Phase B peak 13.1 2θ in the sample 12/30/2009 Richard E. Young 39
  40. 40. PXRD RIR Slope Determination for Weight% Ratio Versus Peak Intensity Ratio 2 1.8 Weight % Ratio (XBXA) 1.6 1.4 1.2 1 0.8 0.6 0.4 0.2 0 0 0.1 0.2 0.3 0.4 0.5 0.6 0.7 0.8 0.9 1 2θ Peak-Height Ratio (IB/IA ) (XBXA) = 1.976 × (IB/IA) – 0.0013 r2 = 0.991 12/30/2009 Richard E. Young 40
  41. 41. ATR-FTIR Spectra of Phase A, Phase B, and Placebo Placebo Phase A Phase B 12/30/2009 Richard E. Young 41
  42. 42. FT-Raman Spectra of Phase A, Phase B, and Placebo Placebo Phase A Phase B 12/30/2009 Richard E. Young 42
  43. 43. ATR-FTIR and FT-Raman Polymorph Calibrations Calculated vs Ac tual Plot - Phas e B 51 Corr. Coeff.: 0.99445 RM SEC: 1.54 ATR-FTIR r2 = 0.989 Ca lcula te d Calibration Validation Correction -2 -2 Actual 51 C alculated vs Ac tual Plot - Phas e B 51 C orr. C oeff.: 0.98488 R M SEC : 2.53 FT-Raman r2 = 0.970 Ca lcula te d C alibration Validation C orrection -2 -2 Actual 51 12/30/2009 Richard E. Young 43
  44. 44. Determined Weight% of Phase B by PXRD, ATR-FTIR, and FT-Raman Lot 1: 0 wt% Phase B Sample ID PXRD ATR-FTIR FT-Raman Rep-1 0.0 1.7 2.2 Rep-2 0.0 0.0 -0.10 Rep-3 0.0 1.0 0.7 Rep-4 0.0 2.0 1.8 Rep-5 0.0 2.0 2.4 Mean 0.0 1.3 1.4 Stnd. Dev. 0.000 0.853 1.07 12/30/2009 Richard E. Young 44
  45. 45. Determined Weight% of Phase B by PXRD, ATR-FTIR, and FT-Raman Lot 2: 10 wt% Phase B Sample ID PXRD ATR-FTIR FT-Raman Rep-1 12 12 16 Rep-2 12 12 10 Rep-3 11 14 15 Rep-4 11 15 16 Rep-5 12 14 17 Mean 12 13 15 Stnd. Dev. 0.548 1.34 2.77 12/30/2009 Richard E. Young 45
  46. 46. Summary of Case Study 2 Two Polymorphs of a Research Compound • PXRD produced quantitative results superior to either ATR or Raman. • ATR produced quantitative results superior to Raman • Another chemometric calibration method might work improve the vibrational spectrocopic results 12/30/2009 Richard E. Young 46
  47. 47. Conclusions • All of the Techniques (PXRD, ATR, Raman) are Non-destructive • All of the Techniques can be Applied to APIs as well as Drug Products • The Vibrational Techniques (ATR and FT-Raman) have Demonstrated Capabilities for Quantification of Complex Polymorph Mixtures • The Superiority of a Particular Technique for Quantification is on a Case by Case Basis 12/30/2009 Richard E. Young 47
  48. 48. Acknowledgements • Amid Salari • Kewei Xu • Fujun Li • Tobin Koppelmaa • Lourdes Javier • Lilia Limon 12/30/2009 Richard E. Young 48
  49. 49. Questions 12/30/2009 Richard E. Young 49

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