Concentration
Evaluation of Crystallization and Precipitation in Highly Supersaturated Aqueous Solutions. Lynne S. Taylor Industrial and Physical Pharmacy
Acknowledgements People • • • • • • • • • • • •
Shweta Raina Grace Ilevbare Bernard Van Eerdenbrugh Tian Xie Luis Sousa Anura Indulkar David Alonzo Greg Stephenson Susan Reutzel Edens Gao Yi Geoff Zhang Kevin Edgar and group
Funding • NSF DMR 0804609 • Pfizer Fellowship • Abbvie • Eli Lilly and Company • NSF PFI-AIR 1312157 • DOK-CPPR • NSF 12110140 • NSF ERC EEC-0540855 • Argonne National Lab
Marcus Brewster’s Spring and Parachute Concept
Brouwers J, Brewster ME, Augustijns P. Supersaturating Drug Delivery Systems: The Answer to Solubility-Limited Oral Bioavailability? J Pharm Sci. 2009 98(8):2549-72.
What do we Mean by Supersaturation? activity solid > activity solute
dissolution
𝐶
S= 𝐶 𝑠𝑠
𝑒𝑞
saturated activity solid = activity solute
undersaturated
supersaturated activity solid < activity solute
crystallization
Why are we Interested in Supersaturation?
Increasing supersaturation = increasing flux Davis and Hadgraft Int. J. Pharm. 1991
Supersaturation is not the same as solubility enhancement
Twist and Zatz J. Soc. Cosmet. Chem. 1986
How high can you go? Factors that can limit supersaturation • Crystallization • Release properties of formulation • Solubilization by formulation components • Upper limit of achievable supersaturation dictated by spinodal decomposition (amorphous solubility)
Predicted vs Observed Amorphous:Crystalline Solubility Ratio 40 Predicted Measured
a
c
Solubility Ratio (S /S )
35 30 25 20 15 10 5 0 Indomethacin
Griseofulvin
Hydrochlorothiazide
Hancock, B.C. et. al., Pharmaceutical Research, Vol. 17, No. 4 2000. 8
Influence of supersaturation on nucleation and growth rates Nucleation Rate1 L-Histidine
Growth Rate m-aminobenzoic acid
1Jiang
S, ter Horst JH 2010. Crys. Growth Des. 11(1):256-261.
Growth Rate (g/ml/min)
0.75
0.50
0.25
0.00 0.0
2.5
5.0
7.5
10.0
12.5
Supersaturation Ratio (C/Ceq)
• So theory tells us that if the supersaturation generated by a solubility enhancing formulation is higher, crystallization is more likely to occur
• Particularly relevant for amorphous solids with a high “solubility advantage”
Variation in Crystallization Kinetics of Highly Supersaturated Solutions
Van Eerdenbrugh, et al (2014). Classification of the Crystallization Behavior of Amorphous Active Pharmaceutical Ingredients in Aqueous Environments. Pharm. Res. 31(4):969-982.
Variation in Crystallization Kinetics of Highly Supersaturated Solutions Metastable polymorph 60 min 30 min 5 min 150s Flufenamic Acid
Ketoconazole
Range of Crystallization Rates in Highly Supersaturated Solutions
Total Percentage
Class I (60 min)
4-Biphenylcarboxylic Acid 4-Biphenylmethanol 4-Phenylphenol Benzamide Caffeine Carbamazepine Felbinac Indoprofen Lidocaine Loviride Naproxen Phenacetin Piroxicam Tolazamide Tolfenamic Acid
Aceclofenac Acetaminophen Anthranilic Acid Celecoxib Chlorpropamide Chlorzoxazone Dibucaine Dipyridamole Felodipine Fenofibrate Flufenamic Acid Flurbiprofen Griseofulvin Haloperidol Nilutamide Nimesulide Tolbutamide 17 36
Bifonazole Carvedilol Cinnarizine Clotrimazole Clozapine Efavirenz Ibuprofen Indomethacin Itraconazole Ketoconazole Ketoprofen Loratadine Miconazole Probucol Procaine
15 32
15 32
Some Amorphous Compounds Dissolve to the Predicted Value Prior to Crystallization – This is Rare
Dissolution of Amorphous Felodipine 10
Theoretical Solubility Advantage: ~9 g/mL
9
Concentration (g/mL)
8 7 6 5 4 o
3
37 C Amorphous o 37 C Crystalline
2 1 0 0
10
20
30
40
50
Time (min) Alonzo et. al., Pharmaceutical Research. Vol 27, No. 4, 2010.
60
70
80
90 15
Crystallization Behavior 1 min
3 min
5 min
10 min
15 min
20 min
Amorphous felodipine exposed to pH 6.8 buffer
16
Ability of Polymers to Modify Solution Crystallization Behavior Supersaturated solution
x
Nucleation
x
Crystal Growth
Amorphous Indomethacin Dissolution (25°C) 30
25
Concentration (g/mL)
Theoretical prediction ~21 g/mL 20
15
10
5
0 0
50
100
150
200
250
Time (min) 18
Alonzo et. al., Pharmaceutical Research. Vol 27, No. 4, 2010.
Amorphous Indomethacin Dissolution with Polymer (25°C) 30
25C PVP 25C No polymer
Concentration (g/mL)
25
Theoretical prediction ~21 g/mL
20
15
10
5
0 0
50
100
150
200
250
Time (min) 19
Alonzo et. al., Pharmaceutical Research. Vol 27, No. 4, 2010.
Celecoxib Crystallization Behavior – Induction Time Experiments
0 min
1 min
6 min
2 min 25
HPMCAS
Concentration (ug/ml)
20
15
10
PVP K12
No polymer
5
0 0
60
120
180
240 Time (min)
300
360
420
480
Upper Limit of Achievable Supersaturation? supersaturated (unstable,2 phase)
Slow crystallizer
concentration
LLPS Fast crystallizer
L-L coexistance
time
Metastable boundary
supersaturated (metastable, 1 phase)
Spinodal
(amorphous solubility)
crystallization
Crystal solubility subsaturated (1 phase)
Crystallization Routes for Highly Supersaturated Solutions
LLPS
Nucleation
Nucleation
Highly supersaturated solution Dissolution of Supersaturating System
Crystal Growth
Literature Precedents
Upper Limit of Achievable Supersaturation
liquidus
Temperature
I
II
III
Amorphous Solubility/ LLPS
Crystalline Solubility
binodal
•Region I – No phase separation can occur
spinodal
•Region II – Only crystalline phase can form •Region III – Both phases can occur, depends on thermodynamics and kinetics of either phase separation process
Concentration Brick et al. Langmuir, 19, 6367-6380 (2003)
Partial Phase Diagram For a Slowly Crystallizing System Felodipine Phase Diagram 60 Crystalline Solubility
40 Temp (°C)
Amorphous Solubility
LLPS 20
LLPS w HPMC
Formation of colloidal “droplets” 0 0
2
4
6
8
10
12
Concentration (μg/mL)
Raina PhD thesis 2014
14
16
18
20
Maximum supersaturation ratio (amorphous:crystalline) depends on the concentration where LLPS occurs supersaturated (unstable,2 phase)
Slow crystallizer
LLPS
Spinodal L-L coexistance
concentration
time (amorphous solubility)
crystallization
Crystal solubility subsaturated (1 phase)
Formation and Properties of Droplet Phase
Dissolution of a PVP:Ritonavir 90:10 Dispersion
A scattering phase appears above a certain ritonavir concentration and disappears on dilution levbare GA, et al. Molecular Pharmaceutics. 2013;10(9):3392-403
Size Characteristics
Nanoparticle Tracking Analysis Below LLPS
Above LLPS
Further Evidence for Non-Crystalline Nature Pyrene
Fluorescence Probe Data Drug droplet
Thomas and Kalyanasundaram. JACS. (1977)
Crystal
Ritonavir Droplet Phase is a Supercooled Liquid
Temperature (oC)
DSC thermogram of pellet obtained by centrifugation
Precipitate
Supernatant (µg/mL)
Drug
Water
10
95.97 %
4.03 ± 0.53 %
57.1 ± 1.09
37
95.13 %
4.87 ± 1.20 %
36.0 ± 0.49
50
94.85 %
5.15 ± 2.12 %
38.9 ± 0.35
levbare GA, et al. Molecular Pharmaceutics. 2013;10(9):3392-403
32
Crystallization of Two Phase Systems 5 min
30 min
60 min
90 min
120 min
480 min
Samples extracted after dissolution of ritonavir/PVP 10:90 wt. % amorphous solid dispersion.
levbare GA, et al. Molecular Pharmaceutics. 2013;10(9):3392-403
Supersaturation in Solutions with Nanodroplets Diffusion Cell
Temperature Controlled Jacketed Chamber
t
UV Dip Probe Controlled Stirring
Donor 30mL
Receiver Regenerated Cellulose 30mL Membrane 34
Felodipine above LLPS concentration
35
In vivo implications? - Felodipine Flux at 370C
Flux (ug/ml.min)
0.006
0.004 LLPS Onset ~12ug/ml Maximum Activity Maximum Flux
0.002
0 0
20
40
60
80
100
120
Concentration (ug/ml) 6-8K MWCO Membrane in pH 6.8 Raina et al. J. Pharm. Sci. 2014 103(9):2736-48.
36
Explanation
>Amorphous Solubility
LLPS
Supersaturated System
Activity is constant Flux approaches constant, max value
J=
𝐷𝑆𝑎𝑠 ℎ𝛾𝑚 37
Summary • Crystallization from highly dilute, supersaturated aqueous solutions is complex. • Behavior can be substantially modified by additives. • Additive effects are not well understood • Metastable phase transformation, whereby solution is still supersaturated, can occur
Acknowledgements People • • • • • • • • • • • •
Shweta Raina Grace Ilevbare Bernard Van Eerdenbrugh Tian Xie Luis Sousa Anura Indulkar David Alonzo Greg Stephenson Susan Reutzel Edens Gao Yi Geoff Zhang Kevin Edgar and group
Funding • NSF DMR 0804609 • Pfizer Fellowship • Abbvie • Eli Lilly and Company • NSF PFI-AIR 1312157 • DOK-CPPR • NSF 12110140 • NSF ERC EEC-0540855 • Argonne National Lab
Marcus Brewster’s Spring and Parachute Concept
Brouwers J, Brewster ME, Augustijns P. Supersaturating Drug Delivery Systems: The Answer to Solubility-Limited Oral Bioavailability? J Pharm Sci. 2009 98(8):2549-72.
What do we Mean by Supersaturation? activity solid > activity solute
dissolution
𝐶
S= 𝐶 𝑠𝑠
𝑒𝑞
saturated activity solid = activity solute
undersaturated
supersaturated activity solid < activity solute
crystallization
Why are we Interested in Supersaturation?
Increasing supersaturation = increasing flux Davis and Hadgraft Int. J. Pharm. 1991
Supersaturation is not the same as solubility enhancement
Twist and Zatz J. Soc. Cosmet. Chem. 1986
How high can you go? Factors that can limit supersaturation • Crystallization • Release properties of formulation • Solubilization by formulation components • Upper limit of achievable supersaturation dictated by spinodal decomposition (amorphous solubility)
Predicted vs Observed Amorphous:Crystalline Solubility Ratio 40 Predicted Measured
a
c
Solubility Ratio (S /S )
35 30 25 20 15 10 5 0 Indomethacin
Griseofulvin
Hydrochlorothiazide
Hancock, B.C. et. al., Pharmaceutical Research, Vol. 17, No. 4 2000. 8
Influence of supersaturation on nucleation and growth rates Nucleation Rate1 L-Histidine
Growth Rate m-aminobenzoic acid
1Jiang
S, ter Horst JH 2010. Crys. Growth Des. 11(1):256-261.
Growth Rate (g/ml/min)
0.75
0.50
0.25
0.00 0.0
2.5
5.0
7.5
10.0
12.5
Supersaturation Ratio (C/Ceq)
• So theory tells us that if the supersaturation generated by a solubility enhancing formulation is higher, crystallization is more likely to occur
• Particularly relevant for amorphous solids with a high “solubility advantage”
Variation in Crystallization Kinetics of Highly Supersaturated Solutions
Van Eerdenbrugh, et al (2014). Classification of the Crystallization Behavior of Amorphous Active Pharmaceutical Ingredients in Aqueous Environments. Pharm. Res. 31(4):969-982.
Variation in Crystallization Kinetics of Highly Supersaturated Solutions Metastable polymorph 60 min 30 min 5 min 150s Flufenamic Acid
Ketoconazole
Range of Crystallization Rates in Highly Supersaturated Solutions
Total Percentage
Class I (60 min)
4-Biphenylcarboxylic Acid 4-Biphenylmethanol 4-Phenylphenol Benzamide Caffeine Carbamazepine Felbinac Indoprofen Lidocaine Loviride Naproxen Phenacetin Piroxicam Tolazamide Tolfenamic Acid
Aceclofenac Acetaminophen Anthranilic Acid Celecoxib Chlorpropamide Chlorzoxazone Dibucaine Dipyridamole Felodipine Fenofibrate Flufenamic Acid Flurbiprofen Griseofulvin Haloperidol Nilutamide Nimesulide Tolbutamide 17 36
Bifonazole Carvedilol Cinnarizine Clotrimazole Clozapine Efavirenz Ibuprofen Indomethacin Itraconazole Ketoconazole Ketoprofen Loratadine Miconazole Probucol Procaine
15 32
15 32
Some Amorphous Compounds Dissolve to the Predicted Value Prior to Crystallization – This is Rare
Dissolution of Amorphous Felodipine 10
Theoretical Solubility Advantage: ~9 g/mL
9
Concentration (g/mL)
8 7 6 5 4 o
3
37 C Amorphous o 37 C Crystalline
2 1 0 0
10
20
30
40
50
Time (min) Alonzo et. al., Pharmaceutical Research. Vol 27, No. 4, 2010.
60
70
80
90 15
Crystallization Behavior 1 min
3 min
5 min
10 min
15 min
20 min
Amorphous felodipine exposed to pH 6.8 buffer
16
Ability of Polymers to Modify Solution Crystallization Behavior Supersaturated solution
x
Nucleation
x
Crystal Growth
Amorphous Indomethacin Dissolution (25°C) 30
25
Concentration (g/mL)
Theoretical prediction ~21 g/mL 20
15
10
5
0 0
50
100
150
200
250
Time (min) 18
Alonzo et. al., Pharmaceutical Research. Vol 27, No. 4, 2010.
Amorphous Indomethacin Dissolution with Polymer (25°C) 30
25C PVP 25C No polymer
Concentration (g/mL)
25
Theoretical prediction ~21 g/mL
20
15
10
5
0 0
50
100
150
200
250
Time (min) 19
Alonzo et. al., Pharmaceutical Research. Vol 27, No. 4, 2010.
Celecoxib Crystallization Behavior – Induction Time Experiments
0 min
1 min
6 min
2 min 25
HPMCAS
Concentration (ug/ml)
20
15
10
PVP K12
No polymer
5
0 0
60
120
180
240 Time (min)
300
360
420
480
Upper Limit of Achievable Supersaturation? supersaturated (unstable,2 phase)
Slow crystallizer
concentration
LLPS Fast crystallizer
L-L coexistance
time
Metastable boundary
supersaturated (metastable, 1 phase)
Spinodal
(amorphous solubility)
crystallization
Crystal solubility subsaturated (1 phase)
Crystallization Routes for Highly Supersaturated Solutions
LLPS
Nucleation
Nucleation
Highly supersaturated solution Dissolution of Supersaturating System
Crystal Growth
Literature Precedents
Upper Limit of Achievable Supersaturation
liquidus
Temperature
I
II
III
Amorphous Solubility/ LLPS
Crystalline Solubility
binodal
•Region I – No phase separation can occur
spinodal
•Region II – Only crystalline phase can form •Region III – Both phases can occur, depends on thermodynamics and kinetics of either phase separation process
Concentration Brick et al. Langmuir, 19, 6367-6380 (2003)
Partial Phase Diagram For a Slowly Crystallizing System Felodipine Phase Diagram 60 Crystalline Solubility
40 Temp (°C)
Amorphous Solubility
LLPS 20
LLPS w HPMC
Formation of colloidal “droplets” 0 0
2
4
6
8
10
12
Concentration (μg/mL)
Raina PhD thesis 2014
14
16
18
20
Maximum supersaturation ratio (amorphous:crystalline) depends on the concentration where LLPS occurs supersaturated (unstable,2 phase)
Slow crystallizer
LLPS
Spinodal L-L coexistance
concentration
time (amorphous solubility)
crystallization
Crystal solubility subsaturated (1 phase)
Formation and Properties of Droplet Phase
Dissolution of a PVP:Ritonavir 90:10 Dispersion
A scattering phase appears above a certain ritonavir concentration and disappears on dilution levbare GA, et al. Molecular Pharmaceutics. 2013;10(9):3392-403
Size Characteristics
Nanoparticle Tracking Analysis Below LLPS
Above LLPS
Further Evidence for Non-Crystalline Nature Pyrene
Fluorescence Probe Data Drug droplet
Thomas and Kalyanasundaram. JACS. (1977)
Crystal
Ritonavir Droplet Phase is a Supercooled Liquid
Temperature (oC)
DSC thermogram of pellet obtained by centrifugation
Precipitate
Supernatant (µg/mL)
Drug
Water
10
95.97 %
4.03 ± 0.53 %
57.1 ± 1.09
37
95.13 %
4.87 ± 1.20 %
36.0 ± 0.49
50
94.85 %
5.15 ± 2.12 %
38.9 ± 0.35
levbare GA, et al. Molecular Pharmaceutics. 2013;10(9):3392-403
32
Crystallization of Two Phase Systems 5 min
30 min
60 min
90 min
120 min
480 min
Samples extracted after dissolution of ritonavir/PVP 10:90 wt. % amorphous solid dispersion.
levbare GA, et al. Molecular Pharmaceutics. 2013;10(9):3392-403
Supersaturation in Solutions with Nanodroplets Diffusion Cell
Temperature Controlled Jacketed Chamber
t
UV Dip Probe Controlled Stirring
Donor 30mL
Receiver Regenerated Cellulose 30mL Membrane 34
Felodipine above LLPS concentration
35
In vivo implications? - Felodipine Flux at 370C
Flux (ug/ml.min)
0.006
0.004 LLPS Onset ~12ug/ml Maximum Activity Maximum Flux
0.002
0 0
20
40
60
80
100
120
Concentration (ug/ml) 6-8K MWCO Membrane in pH 6.8 Raina et al. J. Pharm. Sci. 2014 103(9):2736-48.
36
Explanation
>Amorphous Solubility
LLPS
Supersaturated System
Activity is constant Flux approaches constant, max value
J=
𝐷𝑆𝑎𝑠 ℎ𝛾𝑚 37
Summary • Crystallization from highly dilute, supersaturated aqueous solutions is complex. • Behavior can be substantially modified by additives. • Additive effects are not well understood • Metastable phase transformation, whereby solution is still supersaturated, can occur
