Selective Production of Polymorphs and Pseudomorphs Using Supercritical Fluid Crystallization from Aqueous Solutions Bouchard,*,†
Jovanovic´,‡
Hofland,†
Andre´anne Natasˇa Gerard W. Eduardo Daan J. A. Crommelin,‡ Wim Jiskoot,‡,| and Geert-Jan Witkamp†
Mendes,§
CRYSTAL GROWTH & DESIGN 2007 VOL. 7, NO. 8 1432-1440
Process Equipment, Delft UniVersity of Technology, Leeghwaterstraat 44, 2628 CA Delft, The Netherlands, Department of Pharmaceutics, Faculty of Pharmaceutical Sciences, Utrecht Institute for Pharmaceutical Sciences, P.O. Box 80.082, 3508 TB, Utrecht, The Netherlands, Section Nanostructured Materials (NSM), Delft UniVersity of Technology, Julianalaan 136, 2628 BL Delft, The Netherlands, and DiVison of Drug DeliVery Technology, Leiden/Amsterdam Center for Drug Research, Leiden UniVersity, P.O. Box 9502, 2300 RA Leiden, The Netherlands ReceiVed NoVember 23, 2006; ReVised Manuscript ReceiVed March 16, 2007
ABSTRACT: A number of supercritical fluid technologies are known to enable the selective production of polymorphs, only by changing the process conditions. These techniques use either supercritical CO2 or organics as solvent. In this work, the precipitation of small organic molecules from aqueous solution was studied using a mixture of supercritical CO2 and ethanol as drying medium and as anti-solvent. Glycine, which has three polymorphs, was precipitated by a direct spraying process. By simple manipulation of the flow rates, the process could be tuned to selectively precipitate either pure R- or β-glycine. When increasing the ethanol concentration in the system, the precipitation of the metastable β-glycine was preferred over the precipitation of R-glycine. Small portions of γ-glycine could be found when choosing slow drying conditions. The same process route was applicable to selectively precipitate pseudomorphs as well. Increasing the ethanol concentration in the extractant phase favored the precipitation of phenylalanine anhydrate over the monohydrate form. The study shows that the supercritical fluid crystallization process has significant potential for the selective production of polymorphs and pseudomorphs of water soluble compounds into small particles in a single step. 1. Introduction Polymorphism plays an important role in the pharmaceutical industry,1 as various polymorphic forms of a chemical compound are being patented for their different physical properties that affect many characteristics such as bioavailability. The polymorphic form is of interest not only for active ingredients but also for excipients as they will influence physical properties of the powder such as its compressibility.2 To date, some pharmaceutical compounds forming polymorphs have been selectively produced using supercritical fluid (SCF) technologies. For example, powders of high polymorphic purity of carbamazepine,3 sulfathiazole,4 and salmeterol xinafoate5 have been produced by selecting appropriate organic solvents, temperature, and pressure while using supercritical (SC) CO2 as anti-solvent. Furthermore, polymorphs of carbamazepine have also been prepared using SC-CO2 as solvent.6 SCF technologies were selected because they allow good control of the precipitation conditions and the use of a single solvent to prepare various polymorphs4 and eliminate a number of steps to produce fine powders of a specific polymorph (i.e., batch crystallization followed by micronization).5 Although water is a common solvent and required for the formation of hydrated pseudomorphs, the selective production of polymorphs and pseudomorphs with CO2 from aqueous solutions has not been previously reported. In the formulation of therapeutic proteins, excipients are generally intended to form an amorphous phase with the proteins, in order to replace the normal aqueous environment of the protein. Yet, situations in which crystalline excipients * Corresponding author. Phone: 31 15 278 5561. Fax: 31 15 278 6975. E-mail:
[email protected] or
[email protected]. † Process Equipment, Delft University of Technology. ‡ Utrecht Institute for Pharmaceutical Sciences. § Section Nanostructured Materials, Delft University of Technology. || Leiden University.
may occur, or are even desired, can be envisioned. For example, glycine and mannitol are common crystalline excipients used to facilitate the drying and improve the cake properties of freezedried formulations of therapeutic proteins,7 and phenylalanine has been reported to reduce the hygroscopicity of formulations.8 The rapid development of protein-based pharmaceuticals encourages the development of alternative drying techniques, such as SCF drying. Appropriate product formulation is also required with these techniques to maintain the long-term protein integrity.9,10 It is then interesting to determine the specific behavior of common excipients in the SCF drying process to gain insight into their crystallization habit once added to formulations and to investigate them as model compounds for more complex active pharmaceutical ingredients in their pure form. SCF technologies have gained attention as a way to obtain micrometer-sized particles of narrow size distribution.11,12 SCF technologies are versatile, as a wide range of supersaturation conditions is attainable through the stepless adaptation of pressure and temperature, next to the normal adjustment of the various flow rates. These conditions can be tuned to achieve a specific density, viscosity, or diffusivity, such that specific interactions with the SCF phase are to favor the precipitation of the preferential polymorph, the crystal habit, etc. Other advantages are: small droplets because of the low surface tension, fast mass transfer, uniform supersaturation conditions, efficient mixing of the solution with the extraction/anti-solvent medium, and quick extraction of the solvent from the particles hindering unwanted recrystallization. CO2 is normally used for such processes, as its critical temperature is close to ambient (Tc ) 31.1 °C, Pc ) 73.8 bar), which also makes it appropriate for the processing of heat labile compounds. Besides, various modifiers (e.g., ethanol, acetone, etc.) can be added to the SCCO2 to tailor the precipitation conditions, especially when water is used as the solvent. The ternary phase diagram of the CO2-
10.1021/cg060834e CCC: $37.00 © 2007 American Chemical Society Published on Web 06/20/2007
Selective Production of Polymorphs and Pseudomorphs
Crystal Growth & Design, Vol. 7, No. 8, 2007 1433
Figure 1. Ternary phase diagram of the CO2-ethanol-water system at 35-40 °C and about 100 bar.13-15
ethanol-water system at about 35-40 °C and 100 bar is given in Figure 1.13-15 Glycine is used as a bulking agent to ease the freeze-drying process of protein formulations.16 Spray-drying of glycine has also been investigated for its potential application in powder formulations for pulmonary delivery.17,18 Glycine has especially interesting characteristics as a model compound for a class of water-soluble pharmaceutical polymorphs. At least three polymorphs of the glycine zwitterion have been described.19-21 β-Glycine, the least stable formsmonotropic transition toward both R- and γ-glycinesis produced under high supersaturation conditions22 and various ways of crystallization have been suggested using ethanol or acetone as anti-solvent.20,23 Dry storage conditions are required to prevent β-glycine from converting to R-glycine.22 R- And γ-glycine are enantiotrops, with γ-glycine being the most stable form at room temperature.24-26 The transformation of γ- to R-glycine happens at 165 °C.21,24 When crystallized from an aqueous solution close to its isoelectric point (pI 5.97), R-glycine is produced.21 γ-Glycine is preferentially crystallized at low and high pH (i.e., below 3.8 and above 8.9).27-29 R-Glycine grows from cyclic dimers,30-32 whereas β- and γ-glycine are built out of monomers. This paper specifies the process conditions required to selectively produce the various polymorphs of glycine by SCF drying. Mannitol, histidine, and cysteine were investigated to verify if the trend observed with glycine could be expanded to other polymorph-forming compounds. L-Phenylalanine was investigated as a model compound for pseudomorphism. The effects of the process conditionssflow rates and temperatures on the pH, polymorph purity, and morphology are discussed. 2. Material and Methods 2.1. Material and Solution Preparation. The following materials were used as received: L-glycine (EP, Fluka), L-phenylalanine (g98%, Aldrich), L-cysteine (g99.5%, Fluka), L-histidine (g99%, SigmaAldrich), D-mannitol (EP, Fluka), ethanol (100%, technical grade), and
CO2 (grade 3.5, Hoek Loos). Aqueous solutions (15-25 g per experiment) used in SCF drying were prepared at least 1 day before drying (at most 1 h before for L-cysteine to avoid the formation of the oxidized cystine form), to ensure complete dissolution. Solutions were prepared with ultrapure water (Maxima HPLC, Elga, Bucks, England) in concentrations appropriate for the compound solubility: glycine, 10% (w/w); phenylalanine, 2% (w/w); cysteine, 5% (w/w); histidine, 2% (w/w); mannitol, 10% (w/w). 2.2. Preparation of Reference Polymorphs and Pseudomorphs. The various polymorphs of glycine were prepared by standard techniques to be used as reference for identification of the SCF dried products. The R- and γ-forms were prepared by freeze-drying of a 5% (w/v) solution.33 For R-glycine, freeze-dried vials containing 1 mL of solution were cooled at a rate of 1 °C/min to -45 °C, kept at that temperature for 5 h, warmed up to -5 °C for 5 h for the annealing step, and finally cooled back to -45 °C at a rate of 0.1 °C/min.33 γ-Glycine was prepared by quench freezing 1 mL volumes of 5% (w/ v) glycine solution with liquid nitrogen in freeze-dried vials. The vials were then put on shelves precooled to -45 °C and kept at that temperature for 4 h. The temperature was then increased to -5 °C for 5 h for the annealing step, and finally decreased back to -45 °C at a rate of 0.1 °C/min.21,33 The pressure was maintained at 0.13 mbar during the entire drying cycle, and vials were stoppered under vacuum. β-Glycine was prepared by anti-solvent precipitation, starting from a saturated solution of glycine in a water-acetic acid mixture (5:1 volume ratio). The filtered solution was mixed with ethanol in a volume ratio of 1:1. The precipitate was recovered by filtration as soon as the solution became turbid. The standard for the anhydrous form of phenylalanine was the purchased material. Phenylalanine monohydrate was produced by cooling crystallization below 37 °C, followed by filtration. The quality of glycine and phenylalanine standards was verified by Raman spectroscopy. 2.3. Experimental Setup, Operating Procedure and Conditions. Aqueous solutions were sprayed into a pressurized precipitation vessel (4-l, 10 cm diameter) together with SC-CO2 enriched with ethanol through a coaxial nozzle. In the experimental setup (Scheme 1), the CO2 was primarily cooled to prevent cavitation, pumped with a diaphragm pump (Lewa), and then heated to the desired drying temperature. The ethanol added from a piston pump (Gilson) was mixed with the SC-CO2 in a T-mixer. This mixture was then directly fed
1434 Crystal Growth & Design, Vol. 7, No. 8, 2007 Scheme 1.
Bouchard et al. Scheme of the Experimental Setup
into the vessel via the outer outlet of a concentric coaxial two-fluid nozzle with inner and outer diameters of 0.15 and 1.1 mm, respectively. The aqueous solution was simultaneously added through the inner outlet of the nozzle using a syringe pump (Isco). Crystallized particles were collected on a filter mounted at the bottom of the vessel. The pressure in the precipitation vessel was controlled using its exit valves. The operating procedure involved the selection of the following process conditions: flow rates, temperature, and pressure. A pressure of 100 bar was selected to minimize the drying time,34 as well as the density fluctuation and inherently the composition, of the CO2-ethanol mixture.35 Once the desired temperature (20, 37, or 70 °C) in the particle-formation vessel and incoming SC-CO2 and ethanol flow rates (4, 7, 10, 13, or 19 mol % ethanol in the SCF phase) were stabilized, the pressurized aqueous solution was sprayed into the vessel. After spraying completion, the vessel was flushed with sufficient SC-CO2 (more than twice the volume of the vessel) to remove the residual water and ethanol from the vessel. Flushing was especially required when the flow rates were such that water was not totally soluble in the SCF phase (water extraction capacity
