Polymorphism of Progesterone - American Chemical Society

Article pubs.acs.org/crystal

Polymorphism of Progesterone: A New Approach for the Formation of Form II and the Relative Stabilities of Form I and Form II Anindita Sarkar, Doaa Ragab, and Sohrab Rohani* Department of Chemical and Biochemical Engineering, The University of Western Ontario, London, Ontario N6A 5B9, Canada S Supporting Information *

ABSTRACT: In the present study, a novel technique has been developed for producing progesterone form II crystals by using shear-assisted sonocrystallization (SAS) method. Progesterone, a steroid hormone, has been recognized for more than 70 years as having two polymorphs, a stable form (form I) and a metastable form (form II). Previous attempts have failed to produce a single crystal of form II of progesterone without the presence of a cocrystal additive or template. The technique proposed in the current study is the first to report the growth of single crystals of progesterone form II. The produced crystals were characterized using X-ray diffraction, differential scanning calorimeter, and Fourier transform infrared spectroscopy. Single crystal X-ray diffraction was performed for comparing the hydrogen bond geometry of forms I and II. Solubility and dissolution rates were estimated, providing insight on the thermodynamics of both forms. Stability studies of both forms were conducted for 60 days, which confirmed the higher stability of progesterone form I. Comparing the crystal structure of form I and form II provides evidence for their relative stabilities. The SAS technique can be proposed as a novel strategy for polymorphic transformation of progesterone, which can increase the dissolution rate, enhance oral bioavailability, and decrease dose-related side effects.

1. INTRODUCTION Progesterone is a poorly water-soluble hormone that belongs to the broad category of progestins (Scheme 1). It is widely

Generally, size reduction is a commonly suggested technique for enhancing the dissolution rate of APIs. However, induced polymorphism, agglomeration, evolution of static charges, and decreased wettability11 are undesirable outcomes of the mechanical process accompanying size reduction. Therefore, isolation of the more soluble polymorph has shown significant promise in enhancing the dissolution rate and solubility of hydrophobic compounds.12 Crystal size, shape, and polymorphic transition are critical factors influencing the physical and chemical properties of the APIs. One of the widely used crystallization techniques in the pharmaceutical industry is antisolvent crystallization.10−14 Numerous antisolvent crystallization systems have been reported for the production of fine quality form I crystals of progesterone. Based on the authors’ knowledge, a single crystal of progesterone form II was only obtained when a structurally related molecule, pregnenolone, was introduced as an additive into the crystallization medium.15 Various methods have been applied to modify the polymorphic forms of inorganic and organic substances.16 Adding template or additive has been proposed as a successful method for polymorphic transformation.17 Dynamic temperature variation (tempering), application of shear, and irradiation

Scheme 1. Chemical Structure of Progesterone

administered as birth control pills and in menopausal hormone replacement therapies.1 Progesterone exists in two monotropically related polymorphs, a thermodynamically stable form I (melting point at 129 °C) and a metastable form II (melting point at 122 °C).2−6 Progesterone can also exist in three other forms (form III, form IV, and form V).7 Yet, to date only polymorphs I and II have been structurally characterized at room temperature and can be found in the Cambridge Structural Database.8,9 A crucial issue associated with poorly soluble drugs is their limited bioavailability, which results from their slow dissolution rate.10 Therefore, various researchers have been focusing on enhancing the solubility, dissolution rate, and oral availability of poorly water-soluble active pharmaceutical ingredients (APIs). © XXXX American Chemical Society

Received: May 7, 2014 Revised: July 30, 2014

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dx.doi.org/10.1021/cg5006727 | Cryst. Growth Des. XXXX, XXX, XXX−XXX

Crystal Growth & Design

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Figure 1. Effect of ultrasonic radiation time on the yield of form II progesterone: (a) 5 min, (b) 20 min, (c) 25 min, and (d) 1 h.

2. EXPERIMENTAL SECTION

with ultrasound waves have been effectively applied to obtain the optimal polymorphic forms of different fats involved in chocolate manufacturing.18−21 Ultrasound-assisted crystallization (sonocrystallization) has recently shown increasing potential in the food and pharmaceutical industry. For the pharmaceutical applications, there are several reports of ultrasound-assisted crystallization.22−24 The detailed mechanism of ultrasound-assisted polymorphic transformation is still unclear. Sonication-induced crystallization has been reported to lead to significantly different properties compared with those produced by simple antisolvent crystallization.25−27 In addition, application of mechanical shear during a crystallization process has resulted in polymorphic transformation.17 Therefore, the current study proposes a new technique for polymorphic transformation of progesterone via a shear-assisted sonocrystallization (SAS) method. Moreover, we were able to grow single crystals of form II progesterone suitable for X-ray diffraction. This was achieved by partially dissolving form II crystals obtained by the shear-assisted sonocrystallization (SAS) method in methanol followed by cooling crystallization leading to the growth of the remaining form II crystals that acted as nuclei. Different solid-state characterization techniques were performed for both crystalline forms of progesterone. Dissolution properties of both polymorphs were investigated, with a focus on the enhanced dissolution rate (DR) of form II.

2.1. Materials. Progesterone (99% pure) was purchased from Calbiochem Company (EMD Biosciences, London, ON) and used without further purification. All solvents used in this work were purchased from VWR Company (VWR International Ltd., London, ON). Laboratory double distilled water was used as an antisolvent. 2.2. Methods. 2.2.1. Preparation of Progesterone Form II Powder Using Shear-Assisted Sonocrystallization Method (SAS). Carefully weighed 0.1 g (3.18 mmol) of progesterone form I was added to 2 mL of methanol at room temperature in a 20 mL vial and exposed to ultrasonic irradiation for 30 min using a Probe Branson Sonifier 450 (Danbery, CT) with 10% of maximum power (400 W), 115 V, and 20 kHz. The sonication phase was followed by insertion of a Polytron PT 2100 Kinematica homogenizer (Luzen, Switzerland) (110 V, 500 W and 50− 60 Hz) in the 20 mL vial for 5 min. Form I (0.1 g) was completely dissolved in 2 mL of methanol (please see the Supporting Information) after applying ultrasonic radiation. Subsequently the solution (2 mL) in the 20 mL vial was added entirely to 100 mL of cold (∼5 °C) deionized water without stirring. A milky solution was formed immediately and highly crystalline materials were formed after keeping the dispersion for 1 day at room temperature. The crystalline material was separated by filtration (5 μm filter paper, VWR brand, 5.5 cm) and dried overnight at room temperature. A yield of 0.086 g (86%) was achieved. 2.2.2. Preparation of Single Crystals Progesterone Form II. In order to grow single crystals of form II, the resulting dry form II crystalline material (0.086 g) obtained by the SAS method was stirred with a magnetic stirrer (magnetic bar, 12 mm × 3 mm) in 0.5 mL of cold methanol (∼5 °C) for 2 min at room temperature (note that according to the solubility curve shown in the Supporting Information, complete dissolution of form II crystals did not happen at room temperature) and B



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Figure 2. Differential scanning calorimetry curves of progesterone form II at different times. Gradual conversion of form II to form I. then kept in a refrigerator (∼5 °C) in a sealed tube. Fine quality needlelike crystals of form II were obtained in about 5−6 h. 2.3. Solid-State Characterization. 2.3.1. Differential Scanning Calorimeter (DSC). The melting points were measured with a Mettler Toledo DSC 822e differential scanning calorimeter (Greifensee, Switzerland). Accurately weighed samples (∼3 mg) were prepared in a covered aluminum crucible having pierced lids to allow escape of volatiles. The sensors and samples were under nitrogen purge during the experiments. The temperature calibration was carried out using the melting point of highly pure indium in the medium temperature range. A heating rate of 5 °C/min was selected. 2.3.2. X-ray Powder Diffraction (XRPD). The XRPD spectra were collected on a Rigaku-Miniflex benchtop X-ray powder diffractometer (Carlsbad, CA) using Cu Kα (λ = 1.54059 Å) radiation obtained at 30 kV and 15 mA. The scans were run from 5.0° to 40.0° 2θ, increasing at a step size of 0.05° 2θ with a counting time of 2 s for each step. The diffractograms were processed using JADE 7.0 software. Calibration was performed using a silicon standard. 2.3.3. Fourier Transform Infrared (FTIR) Spectrometer. FTIR (Bruker, Milton, ON) was employed for quantification of crystal forms. The samples were analyzed in absorption mode through a zinc selenide crystal. Thirty-two scans with a resolution of 2 cm−1 in the range of 600 to 4000 cm−1 were performed. The background was collected in the same range for air. Approximately 2−4 mg of sample was poured on the zinc selenide crystal for each analysis. 2.3.4. X-ray Single Crystal Diffraction. A single crystal of form II of progesterone was mounted on a Mitegen polyimide micromount with a small amount of Paratone N oil. X-ray measurements were made on a

Bruker Kappa Axis Apex2 diffractometer at a temperature of 110 K. The unit cell dimensions were determined from a symmetry constrained fit of 8574 reflections with 4.9° < 2θ < 58.56°. The frame integration was performed using SAINT.28 The resulting raw data were scaled and absorption corrected, using a multiscan averaging of symmetry equivalent data using SADABS.29 The structure was solved using the SIR2011 program.30 Most of the non-hydrogen atoms were obtained from the initial refinement. The remaining atomic positions were obtained from a subsequent difference Fourier map. The hydrogen atoms were introduced at idealized positions and were allowed to ride on the parent atom. The structural model was fit to the data using full matrix least-squares based on F2. The calculated structure factors include corrections for anomalous dispersion from the usual tabulation. The structure was refined using the SHELXL2013 program from the SHELX suite of crystallographic software.31 2.4. Determination of Solubility of Progesterone. The solubility of progesterone in water and in phosphate buffered saline (PBS, pH 7.2) was measured in the range of 21−40 °C. Solubility of form II in methanol was also measured in the temperature range 25−43 °C. To measure the saturation concentration of each form of progesterone, 20 g of the solvent mixture and an excess amount of solids were added to a vial at a given temperature and mixed using a magnetic stirrer plate (AGE Magnetic Stirrer, Newtec Inc., Hull, IA). These suspensions were then placed on a multiplate mechanical shaker and were left to equilibrate for 72 h in a temperature controlled water bath. Samples were filtered through 0.45 μm cellulose acetate syringe filters into volumetric flasks. Supernatants were then analyzed by UV C



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spectrophotometric analysis at λmax 290 nm (Cary 100 Bio UV visible spectrophotometer, Palo Alto, CA). 2.5. Dissolution Tests. In order to evaluate dissolution rate of both polymorphs of progesterone, 100 mg of pure progesterone form I or II was placed in 200 mL of water (25 ± 1 °C) or 100 mL of phosphate buffer saline (PBS, pH 7.2) at 37 °C (representing the simulated body fluid), and the samples were stirred at 100 rpm in a magnetic stirrer for 20 min. Aliquots of 5.0 mL of supernatant were withdrawn every 2 min. The dissolution medium was replaced after every sampling. Dissolution testing was performed in triplicate. The concentration of progesterone in the solution was measured by UV spectrophotometer (Cary 100 Bio UV visible spectrophotometer, Palo Alto, CA). 2.6. Stability Study. Stability studies of form II were performed by storing form II samples in a desiccator with perforated foil at 40 ± 0.5 °C and 75% ± 5% RH for 30 days as per ICH guidelines. The stability of form II and its transformation to form I was determined by the DSC. The stability tests were also performed at 5 °C.

structure of form I crystals matched the previously reported structure (CSD reference PROGST10).15 None of the methods claiming the isolation of form II in the literature resulted in a reproducible recipe. After extensive screening, it was found that though there are a few methods mentioned in the literature for the formation of form II, there is a lack of reproducibility and details in the reported recipes. We conducted the following methods given in the literature with an aim to isolate form II, and the results obtained during our present experimental investigation were invariably form I: (a) Rapid crystallization of crude progesterone (0.31 g, 1 mmol) from acetone solution (5 mL), using a rotary evaporator, as suggested by Wachter,32 resulted in form I. (b) Crystallization of a melt of form I at ca. 125 °C in a DSC pan was also tried, but the outcome was again isolation of form I.33 (c) After equilibrium melting of crude progesterone at 5 °C/min in air at 85 °C for a few hours, the sample was further heated to 122 °C and then slowly cooled to room temperature, and the result was again form I.34 (d) Form I (0.31 g, 1 mmol) was dissolved in 5 mL of ethanol at 40 °C and dried at 50 °C. DSC studies revealed form I with a very small peak of form II.35 (e) Form I progesterone (3.14 g, 10 mmol) was allowed to melt at 150 °C, after which it was poured into 50 mL of deionized water maintained at 5 °C using cryostatic bath, and it was sonicated for 2 min. The process led to the formation of form I.36 (f) Solution of commercial progesterone (form I) in ethanol or n-hexane was evaporated at about 70 °C, and form I was obtained.37 (g) Crystallizing from cold acetone resulted in form I.38 Based on a thorough investigation during our crystallization experiments and allusions to emerging difficulties in the literature, it is tempting to postulate that the growth of form II is problematic and has not been performed successfully. In the light of these failures to obtain form II without pregnenolone as an additive, this study shows a new technique to produce pure progesterone form II by shear-assisted sonocrystallization followed by addition of water as an antisolvent. Ultrasound irradiation time had a strong effect on the yield of form II progesterone in our experiments. Varying the ultrasonic radiation time from 2 min to 1 h affected the percent yield of form II over a wide range with a maximum of 100% at 30 min irradiation time. After 1 h irradiation time, the product obtained was neither form I nor form II. It is postulated that the excess energy input breaks some of the chemical bonds leading to the formation of an unidentifiable structure. Figure 1 indicates the effect of ultrasonication time on the amount of form II produced. Changing the initial mass of progesterone, for example, starting with 0.2 g of progesterone and dissolving it in the same volume of methanol (2 mL), resulted in the formation of a mixture of form I and form II (ca. 1:1) suggesting that the amount of energy input was not sufficient to lead to the formation of pure form II.

3. RESULTS AND DISCUSSION 3.1. Polymorphic Screening. Crystals of form I with a large block habit were easily obtained from a saturated solution of Table 1. Crystallographic Data of Progesterone Polymorphs polymorph chemical formula formula wt cryst size (mm3) cryst syst space group T, K a, Å b, Å c, Å Z V, Å3 Dcalcd, g cm−3 μ [mm−1] reflns collected unique reflns reflns I ≥ 2σI R1 [I > 2σ(I)] wR2 [all] GOF CCDC no. a

form Ia

form II

form II by cocrystallizationa

C21H30O2

C21H30O2

C21H30O2

314.45 0.63 × 0.46 × 0.36 orthorhombic P212121 150(2) 10.2496(7) 12.4830(9) 13.6406(9) 4 1745.3(2) 1.197 0.075 15572 2424 2354 0.0387 0.0976 1.060 PROGST12

314.45 0.41 × 0.06 × 0.027 orthorhombic P212121 110(2) 6.2062(18) 12.573(5) 22.120(10) 4 1726.0(11) 1.210 0.075 25824 5145 3526 0.0498 0.0942 0.9890 CCDC 988960

314.45

orthorhombic P212121 150(2) 6.2089(6) 12.5804(12) 22.188(2) 4 1733.115 1.205 0.075

0.0454

PROGST13

Reference 15.

acetone or methanol or hexane or chloroform by slow evaporation crystallization. The unit cell determination of the

Table 2. Hydrogen Bonding Geometry of Progesterone Polymorphsa D−H···A

d(D−H)

d(H···A)

d(D···A)

D−H···A

2.44(2) 2.49(2) 2.53(3)

3.425(2) 3.455(2) 3.442(3)

155.0(16) 169.4(17) 165(2)

2.56(2)

3.471(3)

155.3(17)

form Ib C(6)−H(6B)···O(3)§1 C(19)−H(19C)···O(3)§2 C(21)−H(21A)···O(3)§3

1.05(2) 0.98(2) 0.93(3) form IIc

C(6)−H(6B)···O(3)§1

0.98(2)

a d in Å and angle in deg. bSymmetry transformations used to generate equivalent atoms: (§1) 1 − x, −1/2 + y, 1/2 − z; (§2) 1 − x, −1/2 + y, 1/2 − z; (§3) 1 − x, −1/2 + y, 1/2 − z. cSymmetry transformations used to generate equivalent atoms: (§1) 1 − x, 1/2 + y, 1/2 − z.

D



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Figure 3. Structure of progesterone form II with the atom labeling scheme. All non-hydrogen atoms are represented by their 50% probability thermal ellipsoids. The torsion angle C16−C17···C20−C21 for form II is shown as 160.39°.

Figure 4. Hydrogen-bonding motifs in the crystal structures of (a) form I and (b) form II of progesterone.

Figure 5. XPRD pattern of progesterone (a) form I and (b) form II.

suspension for 2 min at room temperature using a magnetic stirrer, and placing the suspension in a refrigerator for 5−6 h. Note that the solubility of form II in methanol over the temperature range 5−34.5 °C is less than 0.086 g/0.5 mL which

Fine quality form II crystals with needle-like habit were obtained by partially dissolving 0.086 g of highly crystalline form II powder obtained by the shear-assisted sonocrystallization (SAS) method in 0.5 mL of cold methanol at 5 °C, stirring the E



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showed a very small peak at ∼128 °C, corresponding to the melting point of form I, in addition to the form II peak at ∼122 °C. After 60 days, DSC curves showed that 5% of form II had been transformed into form I. 3.3. Comparison of Crystal Structures of Forms I and II. The comparison of crystal structures of forms I and II provides evidence of their relative stabilities. The single crystal structure of form II obtained in our study was compared with the form II data given by Lancaster et al.15 (PROGST12). The crystallographic data are summarized in Table 1. The hydrogen-bonding geometry is listed in Table 2. The low temperature structure determination, with refined hydrogen atom positions of forms I and II, shows conformational differences in the acetyl group. The conformation of the acetyl group is the key discriminator in determining the packing differences between the two polymorphs of progesterone. The torsion angle C16−C17···C20−C21 varies between 173.10(2)° for progesterone form I and 160.39(2)° in form II. The atomicnumbering of a molecule of form II is depicted in Figure 3. The hydrogen bond that plays an important role in determining crystal packing is different in both forms. Form II of progesterone has only one potential site for hydrogen-bonding interaction, O3 of the oxo group of cyclohexane that participates in intermolecular C6−H6B···O3 hydrogen-bonding interaction. Self-assembly via this C−H···O interaction leads to a onedimensional supramolecular zigzag chain, running parallel to the c axis, in the form II crystal lattice (Figure 4). The oxygen atom in the acetyl group shows a very week C−H···O interaction since the C16···O20 distance and C16−H16···O20 angle are 3.471(3) Å and 104°, respectively. The crystal packing of form I of progesterone is more complicated than that of form II. O3 of the oxo group of the cyclohexane group of one molecule of form I participates in three different intermolecular hydrogen bonding interactions, C6−H6B···O3, C19−H19C···O3, and C21− H21C···O3, with two different packings. All these three interactions lead to a two-dimensional structure in the form I lattice. Again, the acetyl oxygen atom shows a very week C−H··· O interaction in the form I lattice due to a very small C16− H16B···O20 angle of 105.3°. The crystal packing of form I is illustrated in Figure 4a. Therefore, we can conclude that form I, due to its more complicated packing in crystal lattice, is more stable than form II. This is consistent with our experimental observations showing the transformation of form II to form I. The investigation of crystal structure provides positive support with respect to the low solubility of form I in water because the molecule of form I is firmly attached in the lattice due to more complicated hydrogen-bonding interactions. 3.4. X-ray Powder Diffraction. The XRPD patterns of pure samples of progesterone forms-I and II were analyzed and compared with the Cambridge Structural Database (CSD) crystallographic data (PROGST12, form I)15 and the simulated pattern of form II, obtained in the present study. Subsequently, these patterns were used as references to analyze the samples obtained in the crystallization screening study, presented in the following section. Figure 5 shows the simulated and experimental patterns of both progesterone polymorphs. We identified the characteristic peaks of form I to be at 12.95° and those of form II to be at 13.94° and 16.42° 2θ values. The characteristic peaks are listed in Table 3. The XRPD pattern for crystals obtained by the SAS method match form II, and no peak of form I was observed even if the sample was analyzed 60 days after of preparation.

Table 3. XPRD Peaks of Solid Form of Progesterone Polymorphs polymorph form I form II

characteristic peaks 2θ, deg 10.85 10.65

12.95 13.94

14.89 14.79

15.59 16.42

17.21 17.91

22.86 20.03

Table 4. Solubility (mol/L) of Progesterone Solid Forms in Water and in PBS in water temp (°C) 21.0 25.0 30.2 34.5 40.8

form I

in phosphate buffer form II

−6

1.81 × 10 1.84 × 10−6 2.83 × 10−4 3.22 × 10−3 7.20 × 10−3

form I −2

1.04 × 10 1.16 × 10−2 1.48 × 10−2 3.80 × 10−2 3.98 × 10−2

form II −6

2.56 × 10 2.65 × 10−6 4.00 × 10−4 5.78 × 10−3 8.03 × 10−3

3.65 × 10−2 3.79 × 10−2 4.20 × 10−2 5.88 × 10−2 6.30 × 10−2

Table 5. Heat of Solution and the Difference between Gibbs Free Energy of Two Progesterone Forms in Water heat of solution (kJ/mol) ΔHsol1−2 (kJ/mol) ΔG1−2 at 298 K (kJ/mol)

form I

form II

−124 −22 −21

−146

Figure 6. Dissolution profiles of progesterone polymorphs.

indicates that some of the form II powder was not dissolved in 0.5 mL of methanol, and the undissolved form II solids acted as nuclei or seeds for the growth of form II single crystals. The solubility data of form II in methanol at different temperatures are given in the Supporting Information. With the same sonication time, the same amount of antisolvent, and the same rapid cooling crystallization conditions, five different attempts were made, and the product was always form II crystals. Because the amount of form II produced in a single SAS experiment was not sufficient to analyze the thermal behavior, X-ray diffraction, and solubility and dissolution rate studies, we collected the crystalline form II from multiple SAS experimentations. 32. Thermal Analysis. The stability of progesterone form II obtained by our SAS method was evaluated by DSC study (Figure 2). Immediately after the preparation of crystalline form II by the SAS method, the DSC curve showed only one endothermic peak at ∼122 °C corresponding to the melting point of form II. But the same crystalline form II, after 2 days, F



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Figure 7. Differential scanning calorimetric plots of progesterone form II of stored samples at 40 °C for (a) 24, (b) 48, and (c) 60 h and (d) at 5 °C for 80 h.

The XRPD pattern of nondetectable form I diffraction was not unusual because both form I and form II have the same crystal symmetries. This was previously observed in the case of β-Dallose when form I was present as impurity in form II sample and the impurity did not show significant diffraction anywhere in the XRPD pattern of form II.39 We performed a quantitative XRPD method by mixing 5% to 30% of form I as an impurity in pure form II (please see the Supporting Information). For 5% form I, we did not observe any diffraction of form I even at a lower scan rate and smaller step intervals. Therefore, it can be assumed that 5% can be the limit of detection of XRPD of this compound.40 3.4. Fourier Transform Infrared Spectroscopy. The FTIR spectra of progesterone forms I and II showed a distinctive band due to the out-of-plane bending of hydrogen bonded to an sp2 carbon (C2−C4) at 871 cm−1 in form I that was shifted to 862 cm−1 in form II. These results are in agreement with previously reported FTIR values of progesterone in the literature.41−43 Moreover, the stretching peak at C20 of the FTIR is 1700 cm−1 for form I, and a blue shift at 1705 cm−1 of form II was observed. The FTIR spectrum revealed that the SAS method produced form II. FTIR studies also reveal that both of the polymorphs differ with respect to bond length. 3.5. Solubility Data. The results of solubility measurements of forms I and II in deionized water and phosphate buffered saline (PBS, pH 7.2) are given in Table 4, which indicate that form I has a lower solubility than form II and the difference between the solubility of the two forms increases with

temperature. The calibration and validation parameters of UV spectroscopy have been discussed in detail in the Supporting Information. Some other thermodynamic properties can also be calculated from the solubility data. The Van’t Hoff equation for an ideal solution shows that the logarithm of mole fraction of a solute is a linear function of heat and entropy of solution: ln x 2 =

ΔHsol ΔSsol + +c RT R

where x2 is the mole fraction of solute in the solvent, ΔHsol is the heat of solution, ΔSsol is the entropy of solution, T is the absolute temperature, R is the universal gas constant and c is a constant. Mole fraction, x2, can be calculated using the solubility data in the following equation: x=

s × MWsolvent 100 × MWsolute

and then, x2 =

x 1−x

where s is the solubility in terms of gram solute per 100 g of solvent and MW is the molecular weight (g/mol). Plotting ln x2 versus reciprocal of absolute temperature should result in a straight line with a slope of ΔHsol/R. The difference between the Gibbs free energy of the two forms, ΔG1−2, can be calculated from the solubility data as is shown below: G


Crystal Growth & Design ΔG1 − 2 ≅ RT ln

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s1 s2

ACKNOWLEDGMENTS We are thankful to Dr. Paul Boyle of Chemistry Department, Western University, for fruitful suggestions regarding single crystal X-ray diffractrometry. We are also thankful to the Natural Sciences and Engineering Research Council of Canada for its financial support of this project through the Discovery Grant.

where s1 and s2 are the solubilities of forms I and II at an absolute temperature T. Table 5 presents the heats of solution of each form of progesterone, calculated from the solubility data of forms I and II in water, the difference between heats of solution, and the Gibbs free energy at 25 °C. All these data support that transformation of form II to form I is possible but the reverse is not. 3.6. Dissolution Rate. Figure 6 shows the dissolution profile of both progesterone polymorphs in water and phosphate buffer pH 7.2. The calculated rate of dissolution of form II in water was 8.12 × 10−3 mg min−1 mL−1, and that of form I was 2.19 × 10−3 mg min−1 mL−1. The calculated rate of dissolution in phosphate buffered saline (PBS, pH 7.2) at 37 °C of forms I and II are higher than those in water, and the values are 1.15 × 10−2 mg min−1 mL−1 and 4.6 × 10−2 mg min−1 mL−1, respectively. 3.7. Stability Study. Figure 7 shows that storing a sample of form II progesterone at 40 °C and 75% ± 5% RH leads to a faster disappearance of the endothermic peak at ∼122 °C of form II and formation of the endothermic peak at ∼128 °C of form I, than storing it at room temperature. Storing form II samples at ∼5 °C shows even slower growth of form I endothermic peak than at room temperature. This stability study reveals that form II can be stored at low temperature to reduce transformation to form I.

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REFERENCES

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4. CONCLUSION In this paper we propose a novel technique to isolate progesterone form II, the metastable form of progesterone that has a higher solubility. The solubility measurements and dissolution rate experiments demonstrate that form II progesterone is more soluble than form I. DSC, PXRD, FTIR, and single crystal structure determinations confirm the formation of form II progesterone by our SAS method. Also the crystal structures of the two forms provide positive support with respect to the low solubility of form I. The molecules of progesterone in form I are firmly attached in the lattice due to more complicated hydrogenbonding interactions. Though slow transformation of form II to form I is observed by DSC studies and confirmed by thermodynamic properties, calculated from solubility data, the rate of transformation can be controlled if form II crystals are stored at low temperature.

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Article

S Supporting Information *

Crystallographic cif files, calibration curve of UV-vis spectroscopy of form I of progesterone, calibration curve of temperature vs. solubility of form II in methanol, and limit of detection (LOD) of form I by XRPD. This material is available free of charge via the Internet at http://pubs.acs.org. Crystallographic data for form II progesterone has been deposited with the Cambridge Crystallographic Data Centre (deposition number is CCDC 988960). Crystallographic information files are available from the Cambridge Crystallographic Data Center (CCDC) upon request (http://www.ccdc.cam.ac.uk). Corresponding Author

*E-mail: [email protected]. Notes

The authors declare no competing financial interest. H



Article

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Polymorphism of Progesterone - American Chemical Society

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