High throughput pre-formulation studies using automated crystallization and analyses

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High throughput pre-formulation studies using
automated crystallization and analyses
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Pre-formulation Workflow at Symyx
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Pre-formulations Discovery Tool System
EpochTM
EpochTM
Library

Studio

Library Studio
Spectra Studio TM
Sample Preparation Station
Birefringence Station
Impressionist
Raman Station
EpochTM
Universal Substrate TM
EpochTM
Crystallization Station
Spectra Studio TM
Polyview
Polyview
XRD Station
EpochTM
EpochTM
U.S. Patents No. 6157449 6371640, and
6004617. Additional Patents Pending
Solubility Station
Melting Point Station
Renaissance Software Suite Integration
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Symyx crystallizer assembly
Block with channels
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Procedure for Typical Automated Crystallizations

• Dispense a solution or slurry of API (as well as
  acid or base solutions for salt selection) to the
  slurry crystallizer (typically 5 to 10 mg of API
  in 200 uL/well) using liquid handling robot at
  Sample Preparation station
• Evaporate dispensing solvent
• Bring crystallizers to Crystallization Station
• Use Epoch software to execute the following
  steps
• add recrystallization solvents to the slurry
  crystallizer (800 uL) and anti solvents to the
  precipitation crystallizer (300 uL)
• equilibrate slurry cyrstallizer at Thigh for 2
  hours
• filter at Thigh and dispense aliquots of the hot
  filtrates to
• evaporation crystallizer at RT (200 uL)
• precipitation crystallizer at RT (100 uL to 300
  uL anti-solvent)
• controlled cooling crystallizer at Thigh (200 uL)
• A plate of vials for HPLC measurement at Thigh
  (50uL)
• execute controlled cooling cycle (Thigh to Tlow,
  8 hours)
• Transfer aliquots from controlled cooling
  crystallizer after cooling cycle for HPLC
  measurement at Tlow (50uL)
• remove solvents from crystallizers
• Open crystallizers and dry crystals
• Isolate Universal Substrate from crystallizer and
  characterize solids

Cycle time 1 day
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Flow Chart for Typical Automated Crystallizations
Heated Filter plate
Slurry API (5 - 10 mg/well) Solvents (800
uL/well)
650 uL
200 uL
100 uL
200 uL
50 uL
Evaporation
Cooling
LC hot
Precipitation
50 uL
LC cold
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Crystallization station
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Core Module
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Birefringence Station
Evaporation
Slurry
Cooling
Precipitation

• Epoch
• Controls automated acquisition
• translates through the library
• auto focuses on each well
• captures a polarized image
• stores images to the database

144979 slurry F8
144982 cooling D11
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Raman Station
Spectra Studio TM Allows user to define a
correlation factor and sort spectra into
groups. to overlay multiple spectra for
comparison. Visualizes groups in library format.
Epoch Allows user to map crystals for Raman
analysis and stores images and coordinates to the
database. Automatically collects spectra for each
position and stores acquisition parameters and
data to the database.
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XRD Station
Spectra Studio TM
Epoch TM Allows user to map positions for XRD
data acquisition and stores images and
coordinates to the database. Automatically
collects data for each position and stores the
acquisition parameters, area plots, and 2q plots
to the database.
Universal Substrate
U.S. Patent No. 6317640. Additional Patents
Pending
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Parallel Melting Point Station
Epoch Allows user to set initial and final
temperature (e.g. 40oC to 300oC) and ramp rate
(e.g 1oC/minute). Controls temperature and
automatically collects data (the intensity of
polarized light) as a function of
temperature. Specifications 4
hours/plate Precision lt0.5oC, Accuracy lt3.0oC
Mixture of Polymorph I and II
intensity
Polymorph II MP65C
e.g. Polymorph screening of Nabumetone
Polyview Allows user to view traces of
intensity versus temperature stored on the
database. to view transition temperatures. to
export results to third party software, e.g.
Excel, Spotfire, etc.
Polymorph I MP80C
temperature
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Solubility Station
Symyx Liquid Handling Robot
Epoch Controls execution of dilution and stores
dilution factor in database. Controls acquisition
of LC data and stores acquisition parameters and
LC data to the database.

• Polyview
• Allows user
• to view LC traces of intensity versus retention
  time and peak areas.
• to export results to third party software, e.g.
  Excel, Spotfire, etc.

Agilent LC
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Renaissance Data Browser
Project Name (Library List)
Birefringence Exp (Images)
Synthesis Exp (Composition)
Library (Experiment List)
Raman Mapping Exp (Images, Coordinates)
XRD Form Exp (Assignments)
Solubility Exp (Values, Traces)
XRD Exp (2q Plots)
Raman Exp (Spectra)
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Capabilities

• Research as of 3/15/2004
• Collaborations with Pharmaceutical Companies 18
• Pharmaceutically Active Compounds 80
• Crystallizations 95,000
• Crystalline Forms Found 470
• FTE Year 10
• Pre-formulation Discovery Tool Capacity
• APIs/year 50
• Crystallizations/Year 50,000
• FTE 5

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General Library Designs
Salt Selection Designs compositional diversity is
generate by varying counter-ion in one dimension
and crystallization solvent in the other
Polymorph Designs Diversity is created by using
different solvents and binary solvent mixtures
created by dispensing gradients of the two
solvents, e.g. four main solvent and 16
co-solvents give 84 unique solvent compositions.
European Patent EP1080435 Additional Patents
Pending
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Library Design Solvent Selection
N8
1
2
3
4
5
6
7
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Solvent Hierarchy Clusters Solvents (1 lt n lt 108)
Solvent Properties Table 108 Solvents 15 Physical
Properties
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Case study
Salt selection using core module with 72 well
Symyx crystallizer assembly
Ephedrine
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Design 14005
None Acetic acid HBr HCl Citric acid Maleic
acid Methanesulfonic acid Phosphoric
acid Sulfuric acid Tannic acid L-tartaric
acid p-toluenesulfonic acid
Salt selection procedure 1. Dispense solution
of API (e.g. methanol, 10 mg/well) 2. Dispense
solutions of acids (e.g. methanol) 3. Equilibrate
(e.g. 50oC 2 hrs) 4. Remove reaction solvent 5.
Crystallization (800 uL/well)
Water Ethanol Acetonitrile 2-butanone 1,4-dioxane
Trifluorotoluene
Filter plate
650uL
Slurry 144503
50uL
200uL
100uL
200uL
Evaporation 144504
Cooling 144506
LC hot (144503)
Precipitation 144505
Cooling 65 to 10ºC 8 hours
Precipitate with 300uL anti-solvents
50uL after cooling cycle
LC cold (144506)
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Design 14005 Birefringence (2.5x)
slurry
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Design 14005 Raman Sorting
slurry
1
11
2
3
4
5
6
7
8
9
10
12
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Design 14005 Raman Comparison
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11
10
9
8
7
6
5
4
3
2
1
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Design 14005 XRD Sorting
slurry
2
3
4
5
6
7
8
9
10
12
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Design 14005 XRD Comparison
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10
9
8
7
6
5
4
3
2
1
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Ephedrine Maleate anhydrous Form B
R10X10
Birefringence Raman
XRD
R10
X10
DSC TGA NMR
1 eq Maleic acid peak
104oC
1eq Maleic acid, slurry in a,a,a-trifluorotoluene
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Forms found
10 ephedrine salts were found in this single set
of experiments (72x4288 crystallizations)
Assignment Assignment Assignment Compound Counter Ion Counter Ion Cocrystallite Cocrystallite Polymorph
Form Raman XRD Compound name eq name eq Polymorph
R1X1 R1 X1 Ephedrine         A
R2X2 R2 X2 Ephedrine chloride 1.0     A
R3X3 R3 X3 Ephedrine methanesulfonate 1.0     A
R4X4 R4 X4 Ephedrine maleate 1.0     A
R10X10 R10 X10 Ephedrine meleate 1.0     B
R5X5 R5 X5 Ephedrine p-toluenesulfonate 1.0     A
R6X6 R6 X6 Ephedrine bromide 1.0     A
R7X7 R7 X7 Ephedrine phosphate 0.5     A
R8X8 R8 X8 Ephedrine sulfate 1.0 water 1.0 A
R12X12 R12 X12 Ephedrine L-tartrate 0.5 water 1.0 A
R9X9 R9 X9 Ephedrine L-tartrate 1.0     A
 R11 R11 uncharacterized   uncharacterized   uncharacterized   uncharacterized   uncharacterized   uncharacterized  
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Form assignment
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Form assignment of Design 14005
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Acknowledgements

• Engineering
• William Chandler
• Chris Freitag
• Trevor Frank
• John Galipeo
• Dennis Mullins
• James Bennet
• Tuyen Nguyen
• Software
• Henry Chau
• Tom Crevier
• Rolland Lou
• Kim Phan
• Jon Rogers
• Dave Dorsett
• John McCarthy
• David Pirkle

• Analytical
• Peijun Cong
• Robert Doolen
• Shan Lin
• Li Song
• Chemistry
• Eric Carlson
• Peter Desrosiers
• Colin Masui
• Anny Tangkilisan
• Quyen Ung
• Eva Wu
• Jessica Zhang
• Thomas Kudla
• Laurie Hall
• Helming Tan
• Fereshteh Lesani
• Isabel Galdo
• Jonah Troth