Elucidating the Complex Phase Behavior of a Cocrystal System

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Elucidating the Complex Phase Behavior of a Cocrystal System Containing Two APIs and One Coformer Paulene Maria Abundo, Zai-Qun Yu, Pui Shan Chow, and Reginald B. H. Tan Cryst. Growth Des., Just Accepted Manuscript • DOI: 10.1021/acs.cgd.8b01238 • Publication Date (Web): 28 Nov 2018 Downloaded from http://pubs.acs.org on November 29, 2018

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Crystal Growth & Design

Elucidating the Complex Phase Behavior of a Cocrystal System Containing Two APIs and One Coformer Maria Paulene Abundo†, Zai-Qun Yu*,†, Pui Shan Chow†, Reginald B. H. Tan*,†,‡ †

Institute of Chemical& Engineering Sciences, A*STAR (Agency for Science, Technology and Research), 1 Pesek Road, Jurong Island, Singapore 627833 ‡

Department of Chemical & Biomolecular Engineering, National University of Singapore, 10 Kent Ridge Crescent, Singapore 119260

Emails: [email protected], [email protected]. Abstract A method has been developed to determine the phase behavior of a cocrystal system that contains two APIs (active pharmaceutical ingredients) and one coformer. Isoniazid, nicotinamide and succinic acid (denoted as A, B and C respectively hereafter) can form a ternary cocrystal ABC, along with two binary cocrystals, A2C and B2C, making the phase behavior very complex because of a variety of eutectic points. The phase diagram of such a complex system has never been reported. During experiment, solids of A, B, C or A2C were added in lots to suspensions of pure ABC cocrystal or a mixture of ABC and another solid phase. Attenuated Total Reflectance Fourier Transform Spectroscopy (ATR-FTIR) was used to measure the concentrations of A, B and C during the process. The constitution of solid phases in suspension at each equilibrium point was determined in a real-time manner by performing mass balance between the liquid and solid phase. Multiple eutectic points in equilibrium with a mixture of ABC and A2C, a mixture of ABC and B2C and a mixture of ABC and A, respectively, were obtained. In the 3D phase diagram spanned by the concentrations of A, B and C, these eutectic points constitute the boundary for a surface where pure ternary cocrystal can be obtained. Eutectic points in equilibrium with three solid phases simultaneously were not reached. With such a phase diagram, a cocrystallisation process can be designed that thermodynamically guarantees the production of pure ABC cocrystal. In addition, practical issues and resolutions in applying ATR-FTIR to phase diagram measurement were discussed. Key words: ternary cocrystal, cocrystallisation, phase diagram, ATR-FTIR 1 ACS Paragon Plus Environment

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Introduction

The ability of cocrystals to confer active pharmaceutical ingredients (APIs) with superior physiochemical and pharmacokinetic properties through improvements in solubility, bioavailability, dissolution and stability, has been widely reported.

1,2,3

These advantages have

spurred the discovery of binary cocrystals in the last decade, such as isoniazid,

4

danazol 5,

pyrazinamide 6 and combination drugs. 7 Today, there exists a wealth of information on protocols for binary cocrystal screening 8 and process development, 9 making it one of the most well studied alternative solid forms of API. In contrast, the field of ternary (and higher order) cocrystallisation remains largely unexplored. Design strategies for ternary cocrystals have only recently emerged due to a growing interest in multidrug cocrystals that can eliminate the segregation of different components during manufacture. 10,11,12,13 Compared to binary cocrystallisation, the assembly of ternary cocrystals is less straightforward due to an increasingly complex hierarchy of intermolecular interactions and spatial factors such as molecular packing, shape and size

14,15

that have to be simultaneously optimized within the

lattice. To a large extent, this supramolecular challenge has already been addressed by ternary cocrystal design strategies for a variety of API and coformer functional group pairings as shown in the references cited above. Successful execution of these approaches however, is limited by the fact that ternary cocrystals are not always the sole products of three-solute cocrystallisation attempts. Depending on the system and its starting conditions, single-component crystals and/or binary cocrystals can form instead of, or alongside the desired product, making pure ternary cocrystals elusive and difficult to isolate. This presents a very interesting problem from the perspective of general cocrystal screening and process development. How to design the screening conditions so that a ternary cocrystal will not be missed if there is? How to develop a process that is capable of producing ternary cocrystal consistently? Similar issues have been encountered in binary cocrystallisation, albeit much easier to overcome due to the presence of fewer components and side products. This is often possible through the use 2 ACS Paragon Plus Environment

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of solubility phase diagrams, which provides a means of identifying the cocrystal operating region in order to avoid conditions that favour single-component outcomes 16. The embedded solubility information also allows the control of supersaturation, which can lead to improved crystallisation results 17. Yu et. al., for instance, was able to determine and control the cooling trajectories for the pure binary cocrystal production of caffeine and glutaric acid, resulting in a narrower particle size distribution and better polymorphic control 18. Phase diagrams were also used by Harmsen and Leyssens

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to assess the plausibility of crystallisation-driven chiral resolution of enantiomeric

ibuprofen, and by Chiarella et. al. 20 to explain the occasional failure of certain binary cocrystal screening methods and protocols. All of this demonstrates the invaluable contribution of phase diagrams in advancing binary cocrystallisation screening and process development. Phase diagrams for ternary pharmaceutical cocrystals have never been reported. In a ternary cocrystal system, one or more binary cocrystal can form between constituent components. For example, isoniazid (denoted as A hereafter) and succinic acid (denoted as C hereafter) can form binary cocrystal (2:1),

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nicotinamide (B) and succinic acid (denoted as B hereafter) can form

binary cocrystal (2:1), 22 apart from the ternary cocrystal ABC 10. The molecular structures of A, B and C are shown below for easy reference:

Isoniazid (A)

Nicotinamide (B)

Succinic acid (C)

Scheme 1: Molecular structure of components of A, B and C Therefore, there are six solid phases in total that may exist individually or jointly (eutectic points) in the suspension, including A, B, C, A2C, B2C and ABC. These six solid phases have different relative stability and some of them cannot exist in the suspension at the same time, for example, neither the solid mixture of A and C nor the solid mixture of B and C can coexist with ternary 3 ACS Paragon Plus Environment


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cocrystal ABC because the binary cocrystals are more stable than the crystals of single components. However, the permissibility of other combinations is difficult to determine in priori. Eutectic points have to be carefully identified as they delineate the operating boundaries of the desired ternary cocrystal and ultimately, influence how a crystallization process should be controlled. In this work, the phase diagram of ABC cocrystal system will be constructed using in situ Attenuated Total Reflectance–Fourier Transform Infrared (ATR-FTIR) spectroscopy and more methodology details will be presented in the next section. 2

Complexity of ternary cocrystal diagram and measurement method

2.1 Microscopic images of binary and ternary cocrystals In preliminary screening experiments, crystals of A2C, B2C and ABC were obtained by slurry method at room temperature. Polymorphism was not observed for these three cocrystals in this study. Their microscopic images are shown in Figure 1. b

a

c

Figure 1 Microscopic images of A2C (a), B2C (b) and ABC (c) obtained in this study Images of A2C crystals and B2C crystals were taken under SEM (scanning electron microscopy, JSM-6700F). ABC crystals were observed under optical microscope (Olympus BX51). Please note that different scales are used for these three images. A2C crystals look like hexagonal prisms with varying lengths. B2C crystals are elongated in one dimension with a rectangular cross section in other two dimensions. ABC crystals are rod-like, being much larger than the two binary cocrystals. 2.2 Measurement of binary cocrystal diagram

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As a comparison, the general phase diagram of a binary system (as shown in Figure 2) and measurement methodology will be summarized in this section. There are two eutectic points for a given binary cocrystal system at a given temperature. The solid phases at each eutectic point consist of the binary cocrystal and one of the constituent components.

E1 SN [N]

E2

CN CM

SM [M] Figure 2 A generalized solubility curve of 2:1 cocrystal M2N at a certain temperature. [M] and [N] stand for the molar concentration of components M and N respectively. Concentration SM on x-axis and concentration SN on y-axis (open circles) stand for the solubility of component M and N respectively. E1 (open square) denotes the eutectic point where solid-state component N coexists with cocrystal. The concentration of component M at E1 is CM. Likewise, E2 (open square) denotes the eutectic point where solid-state component M coexists with cocrystal. The concentration of component N at E2 is CN. The dashed line has a slope that corresponds to the stoichiometry of cocrystal M2N. The eutectic points were located using the method presented by Yu et. al.

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Take A2C as an

example. To a saturated solution of A2C at 40 oC, an appropriate amount of solid A was added. Solid A would dissolve and combine with component C in the liquid phase to form cocrystal A2C. As a result, concentration of A would increase while that of C would decrease. The suspension was left to stabilize for 60 min for the solution to reach a new equilibrium point. The equilibrium concentrations of A and C were read from ATR-FTIR and one point in the phase diagram was obtained. Then more solid A was added and the process was repeated until the addition of solid A had no effect on the equilibrium concentrations of A and C. This signaled that the eutectic point between the A2C cocrystal and solid A had been reached. In this way, a A-C concentration curve 5 ACS Paragon Plus Environment


that was in equilibrium with A2C was obtained. Similar experiments were conducted to obtain the binary eutectic conditions for A2C + C, B2C + B and B2C + C. 2.3 Measurement of ternary cocrystal diagram For a ternary cocrystal system, the constitution of solid phases at eutectic points will become more varied considering the number of solids phases available. The following ten types of eutectic points may emerge: 

Ternary cocrystal+binary cocrystal: ABC+A2C, ABC+B2C



Ternary cocrystal+crystals of one single component: ABC+A, ABC+B, ABC+C



Ternary cocrystal+binary cocrystal+crystal of single components: ABC+A2C+A, ABC+A2C+C, ABC+B2C+B, ABC+B2C+C



Ternary cocrystal+crystals of two single components: ABC+A+B

Other combinations are believed to be unattainable due to the relative stability of solid phases. For example, ABC+A2C+B and ABC+B2C+A cannot coexist because A2C can combine with B into ternary cocrystal ABC and it is the same case with ABC+B2C+A. ABC+A+C and ABC+B+C is believed to unattainable as well because A and C can combine into A2C, while B and C can combine into B2C. As for the ten types of eutectic points listed above, it remains to be seen which can be obtained in experiments. 2.4 Determination of solid phase constitution at eutectic points As mentioned earlier, a eutectic point in a binary cocrystal system can be approached by adding excess amounts of the corresponding component. Its attainment can be detected by the invariant concentrations as more solids are added. However, this detection method may not work for a ternary cocrystal system because the relative stability of various solid phases is not known a priori. In other words, we do not know how the addition of one solid phase will affect the fate of other solid phases already present in the suspension. Therefore, it is necessary to test the constitution of solid phases during measurement. This can be done by sampling the suspension for X-Ray Powder Diffraction (XRPD) analysis. However, the scheduling of XRPD diffractometer did not match 6 ACS Paragon Plus Environment

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with the time frame of experiments. Therefore, a method based on mass balance was proposed to determine the species of solid phase in a real time manner. During experiments, the mass of solvent and the accumulative mass of solids added to the suspension were recorded. After the concentrations of A, B and C were measured by ATR-FTIR at each equilibrium point, the respective mass of components A, B and C existing in solid phase can be determined by performing mass balance, because the total amount of each component added to the suspension equals to the sum of the amount existing in liquid phase and the amount existing in solid phases. The molar ratio of components A, B and C in solid phases can be calculated. If the resulting molar ratio complies with 1:1:1 (A:B:C), then the solid phase can be deemed as consisting of ternary cocrystal ABC only. If the molar ratio does not agree with 1:1:1, check if combinations of two or more solids phases can account for it. For example, if combination of 1:1:1 (A:B:C) and 2:1 (A:C) can explain the resulting molar ratio, then the solid phase is deemed as consisting of ternary cocrystal ABC and binary cocrystal A2C. This method to determine the constitution of solid phase at eutectic points was verified by off-line XRPD analysis. In order to cover the phase diagram adequately and to determine its boundaries, solids of A, B, C and binary cocrystal will be added to suspensions. The addition of different solid phases will shift the equilibrium toward different direction in the phase diagram. Following this, the solution was momentarily heated to a slightly higher temperature to facilitate dissolution of the added solids and to prevent fast accumulation of supersaturation. Then the temperature was slowly lowered to 40 °C for stabilization and measurement of equilibrium concentration. Sonication was activated to promote precipitation during the process. After each addition of solids, the concentrations of A, B and C measured by ATR-FTIR were recorded as a point in the phase diagram. 3

Experimental

3.1 Chemicals Nicotinamide and Succinic Acid with a purity of 99% was obtained from Alfa Aesar, Isoniazid with a purity of 99% was obtained from Sigma Aldrich and HPLC-grade Ethanol with 99.7% 7 ACS Paragon Plus Environment


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purity was obtained from VWR Chemicals. All of the chemicals purchased were used as delivered. 3.2 Experimental Set-Up Figure 3 is a diagram of the experimental setup. All crystallization experiments were conducted in a 500 mL flat-bottomed glass crystallizer equipped with an overhead stirrer rotating at 400 rpm. Temperature in the vessel was controlled by a programmable circulator (Julabo FP50-HL). Absorbance spectra were collected using a Nicolet 4700 spectrophotometer (Nicolet Instrument Co.) fitted with a Dipper-210 ATR immersion probe (Axiom Analytical Inc.). Each spectrum had a resolution of 4 cm-1 and was the average of 64 scans in the 600 to 4000 cm-1 range.

Figure 3 Experimental setup (1) ATR-FTIR probe (2) Thermocouple (3) Overhead stirrer (4) Ultrasonicator (5) Heating and cooling circulator (6) 500 mL vessel It has been observed that the ATR probe is prone to encrustation when new solid phases are introduced into the suspension. Newly added solid phases dissolve fast and the liquid phase becomes supersaturated with respect to the existing solid phase. For example, when solids of cocrystal A2C are added to a suspension of ternary cocrystal ABC, part of A2C cocrystal will dissolve and renders the liquid phase supersaturated with respect to ternary cocrystal ABC. Subsequent nucleation and crystal growth tend to deposit a layer of cocrystal on the surface of ATR-probe. It was observed that concentration measurements by a fouled ATR probe deviates drastically from the actual values. Therefore, a 750 W, 20 kHz ultrasonicator (Sonics & Materials Inc. VCX750) was used to promote nucleation in positions away from the ATR-probe. It was 8 ACS Paragon Plus Environment

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operated at 20% power to prevent encrustation on the ATR-FTIR probe during the experiments. Ultrasonicator pulse was applied every 20 s and each pulse lasted for 10 s. 3.3 ATR-FTIR Calibration ATR-FTIR was calibrated following the procedure developed by Togkalidou et. al. 23 80 standard solutions with varying concentrations of isoniazid (A), nicotinamide (B) and succinic acid (C) were prepared, each utilizing approximately 220 g of ethanol as solvent. The solution was then heated to dissolve all solids, followed by cooling at 1 °C/min while ATR-FTIR spectra and temperature data were collected concurrently every 2 min until nucleation took place. A total of 251 spectra were acquired spanning temperatures of 10–58 °C. Principal component regression was used to correlate the solute concentration with the temperature and absorbance in the 800– 1800 cm-1 range. The prediction errors for isoniazid, nicotinamide and succinic acid are 1×10-3 g g-solvent -1, 7×10-4 g g-solvent-1 and 8×10-4 g g-solvent-1 respectively. This level of accuracy is sufficient for engineering purposes. In preparing standard solutions, the amounts of A, B and C added to solvent was determined according to 63 factorial design (3 factors, each factor has 6 levels). The targeted concentration range of A, B and C was from 0 to 0.04, from 0 to 0.07 and from 0 to 0.1 g/g-solvent, respectively. There are 216 treatments in this design. However, the solids added in some of these treatments could not be dissolved completely at the maximum allowable temperature (58 °C in this study to prevent significant loss of solvent during measurement). These treatments were discarded for calibration. Consequently, only 80 treatments in the 63 design were used for calibration. 3.4 XRPD analysis D8 Advance powder X-ray diffractometer (Bruker AXS GmbH, Germany) with Cu-Kα radiation (λ = 1.54056 A) was used for XRPD analysis. The voltage and current applied were 35 kV and 40 mA, respectively. The samples were scanned within a range of 2-50° (2θ) at a scan rate of 2°C/min. 9 ACS Paragon Plus Environment


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Results and Discussion

4.1 Concentration profiles in a typical measurement cycle A measurement cycle started from a suspension with known species of solid phases, such as pure ABC, or a mixture of ABC and A2C, etc. Solids of A, B, C, or A2C were added in lots to shift the system from one equilibrium point to another. After each addition, the suspension was stirred for around 60 min at 40 °C to achieve equilibrium between liquid and solid phases. The amount of solids added should cause significant changes in concentration of at least one of the components. Multiple equilibrium points could be measured in one batch. When the crystallizer was crowded with solids, it was emptied, cleaned and dried for the next cycle.

[A]

0.070

50

[B]

45

0.060 0.050

40

0.040 35 0.030

Temperature (oC)

Concentration (g/g-solvent)

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60

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30

0.020

t2

t1

0.010 0

50

100 150 Time (min)

t3 200

25 250

Figure 4 Typical concentration profiles of isoniazid (A), nicotinamide (B) and succinic acid (C) during the cocrystal solubility experiments. The black dashed lines (e.g, at time t1) denote the addition of A2C. Following the addition of solids, the solution is momentarily heated to speed up dissolution as shown in the embedded temperature profile. The two magenta dash-dot lines indicate the arrival at an ABC+A2C eutectic point. Figure 4 shows the temperature and concentration profiles of isoniazid (A), nicotinamide (B) and succinic acid (C) recorded via ATR-FTIR during a measurement cycle. At t = 0, a suspension of ternary cocrystal ABC at 40 oC was prepared and allowed to equilibrate. A2C crystals were then added into the crystallizer at t = t1 and the solution was momentarily heated to facilitate the 10 ACS Paragon Plus Environment

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dissolution of solids. The solution was then gradually cooled back to 40 oC, causing the system to be supersaturated with respect to ABC. Under sonication, cocrystallisation of ABC cocrystal happened as shown by the simultaneous proportional decrease in concentrations of all three components. Further addition of A2C solids as indicated by the dashed lines led to the production of more ABC cocrystals until t=t2, when excess A2C no longer dissolved and the concentrations of A, B and C did not change any more with the addition of A2C solids. At this point, A2C and ABC cocrystals coexisted in solution, indicating the achievement of a eutectic point. Next, changes in concentrations of A, B and C during solids additions will be analyzed to understand the relative stability of different solid phases. 4.2 Addition of solids B to a suspension of pure cocrystal ABC A suspension of pure cocrystal ABC was prepared and B solids were added to it in lots. The changes in equilibrium concentration and constitution are displayed in Figure 5.

Concentration, g/g solvent

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60


0.100 0.080

AB C ABC+ B2C

0.060 0.040

B2 C [A] [B] [C]

0.020 0.000 0

2

4 6 Number of B Additions

8

10

Figure 5 Changes in equilibrium concentration and constitution of solid phases during the addition of solids of B to a suspension. The starting suspension contained pure ABC cocrystal. A mixture of ABC and B2C appeared after the second addition. B2C was the only solid phase in suspension after the 8th addition. Figure 5 shows that concentration of B increased all the time with the addition of B solids, whereas concentration of C decreased all the way. Concentration of A decreased slightly first to 11 ACS Paragon Plus Environment


a minimum and then went up after that. The third addition of B solid seems a reflection point in concentration curves where B2C started appearing in the suspension. Unexpectedly, all ABC crystals in the suspension disassociated after the last addition and a suspension of B2C was obtained at the end. It seems that crystals of B cannot coexist with a mixture of ABC and B2C. The changes in solid phases are in line with those in concentrations of A, B and C. For the first three additions, concentrations of A and C decreased in proportion. It seems that increase in B concentration ‘crowded out’ A and C in the form of ABC ternary cocrystal. With more additions of B solids, component C in liquid phase was consumed in the form of B2C. Cocrystal ABC dissolved to compensate the depletion of C, which caused accumulation of A in liquid phase. 4.3 Addition of C solids to a suspension of pure cocrystal ABC The phenomenon of complete dissolution of cocrystal ABC was not observed during the addition of another component solids, C, to a suspension of pure cocrystal ABC as shown below.


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0.080

AB C

AB C

0.060

[A] [B]

0.040

[C]

0.020 0.000 0

2

4 6 Number of C Addition

8

Figure 6 Changes in equilibrium concentration and constitution of solid phases during the addition of C solids to a suspension. The starting suspension contained pure ABC cocrystal. Pure ABC was obtained in the suspension after the 5th addition. Figure 6 shows that the concentration of C increased continuously with the addition of C solids and the concentrations of A and B decreased monotonically. During the whole process, the solid phase in the suspension contained pure ABC only. All the added C crystals dissolved. Some of components B and C in liquid phase precipitated out as cocrystal ABC during the process. It 12 ACS Paragon Plus Environment

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should be mentioned that the jump in concentration of C at the 5th point was due to the addition of a much larger amount of C solids (around 4 g) than at previous points (around 1 g). No more C crystals were added after the 6th addition because the suspension became very crowded. The eutectic points that are in equilibrium with cocrystal ABC and C crystals were not reached. 4.4 Addition of C solids to a suspension with a mixture of ABC+A C solids were also added to a suspension containing a mixture of ABC and A. The changes in concentrations and solid phase constitution are displayed in Figure 7.


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0.060

ABC +A AB C

0.040

AB C

[A] [B]

0.020

[C]

0.000 0

2 4 Number of C Additions

6

Figure 7 Changes in equilibrium concentration and constitution of solid phases during the addition of solids of C to a suspension. The starting suspension contained a solid mixture of ABC and A. Pure ABC was obtained after the first addition. It can be seen that A crystals in the suspension disappeared after the first addition of C solids and the solid phase in the suspension contained pure ABC only. Crystals of C dissolved and combined with components of A and B in liquid phase to form cocrystal ABC. Depletion of component A in liquid phase prompted the dissolution of A crystals originally in the suspension. This observation confirms the hypothesis that A crystals cannot coexist with C crystals in suspensions of ABC. After the 1st addition of C crystals, the concentration of A did not change much while the concentration B dropped significantly due to the presence of A solids in the starting suspension. After the 2nd addition, concentrations of A and B decreased in parallel due to the stoichiometry in ABC formation. 13 ACS Paragon Plus Environment


4.5 Addition of A solids to a suspension of ABC+B2C When solids of A was added to a suspension of ABC+B2C, B2C dissolved and more ABC crystals were generated as shown in Figure 8. Concentration, g/g solvent

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ABC+ B2C

0.060 0.050

AB C

ABC +A

0.040

[A]

0.030

[B]

0.020

[C]

0.010 0.000 0

2 4 Number of A Additions

6

Figure 8 Changes in equilibrium concentration and constitution of solid phases during the addition of solids of A to a suspension. The starting suspension contained ABC and B2C. Pure ABC was obtained after the 1st addition of A solids. A mixture of ABC and A was obtained after the 4th addition. After the 1st addition of A solids, the solid phase of suspension contained ABC only. This phenomenon is in line with our analysis in Section 2.2 that solids of A, B2C and ABC cannot coexist. With the addition of more A solids, concentration of A increased and the concentrations of B and C decreased slightly in proportion. After the 3rd addition of A solids, the concentrations of A, B and C stabilized. A mixture of ABC+A appeared in the suspension. 4.6 Addition of A2C to a suspension of pure ABC Apart from solids of A, B and C, solids of A2C were added to a suspensions of pure ABC as well to shift to other points in the phase diagram along a path different than by adding A, or B or C. A2C cocrystals were prepared in a separate experiment by mixing appropriate amounts A solids and C solids in ethanol. It can be seen in Figure 9 that the concentration of A went up significantly during the first three additions of A2C, while the concentration of B dropped significantly. At the same time, drops in 14 ACS Paragon Plus Environment

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C concentration were smaller than that in B. The relative amount of changes in concentrations of A, B and C can be explained by the stoichiometry of A2C and ABC. Increase in A concentration was caused by the dissolution of A2C solids. After one unit of A2C solids dissolved, two units of A and one unit of C were carried into the liquid phase. Then one unit of A, one unit of C combined with one unit of existing B in liquid phase to form cocrystal ABC. As a result, B in liquid phase was consumed while there was surplus of A after precipitation of ABC. Meanwhile, increase in A concentration had an effect of ‘crowding out’ other components from liquid phase. Therefore, slightly more C would precipitate out in the form of ABC than was brought into the liquid phase by the dissolution of A2C solids.


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0.040

A BC

0.030

ABC+ A2C

[A] [B]

0.020

[C]

0.010 0.000 0

1

2 3 4 Number of A2C Additions

5

6

Figure 9 Changes in equilibrium concentration and constitution of solid phases during the addition of solids of A2C to a suspension. The starting suspension contained pure ABC cocrystal. A mixture of ABC and A2C was obtained after the 4th addition. After the 5th addition of A2C solids, the concentrations of A, B and C levelled off and the eutectic point in equilibrium with ABC+A2C was reached. 4.7 Verification of solid phase constitution by XRPD analysis XRPD patterns of samples containing pure A, B, C, A2C, B2C and ABC respectively collected at room temperature are shown in Figure 10 together with the simulated XPRD pattern of ABC.10 The peak shifts towards larger 2-Theta angles between simulated and measured XRPD patterns



of ABC are due to anisotropic expansion of unit cells (single-crystal data of ABC were collected at 110 K while the XRPD pattern was collected at room temperature).

ABC simulated

Relative intensity

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60

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ABC ABC+B2C ABC+A2C ABC+A B2C A2C Succinic acid (C) Nicotinamide (B) Isoniazid (A) 5

15

2Theta,°

25

35

Figure 10 The XRPD patterns of samples containing pure A, B, C, A2C and B2C respectively are displayed in ascending order starting from bottom. On top is simulated XRPD pattern of ABC, followed by measured XRPD pattern of ABC. The patterns of samples withdrawn at eutectic points (detected by mass balance method) were found to contain characteristic peaks of two solid phases and are labeled as ABC+A, ABC+A2C and ABC+B2C, respectively. Samples of slurry were withdrawn during phase diagram measurements for XRPD analysis. Their patterns were compared with the references to determine their constitution. The sample labeled as ABC+A was collected at the eutectic point of ABC+A (as detected by mass balance calculation, Section 2.3). It can be seen that its pattern indeed contains characteristic peaks of both ABC and A. Similarly, samples labeled as ABC+A2C and ABC+B2C were collected at the eutectic points of ABC+A2C and ABC+B2C, respectively. Their XRPD patterns confirmed the constitution of eutectic points. 4.8 Phase Diagram All equilibrium points obtained in this study are presented in the 3D diagram in Figure 11. A fitting surface for all points is drawn to guide the eye. In this fitting surface concentration of C was correlated with the concentrations of A and B in a polynomial form using a Matlab routine, fit([x,y],z,'poly23'). It can be seen that the fitting surface adequately reflects the relationship 16 ACS Paragon Plus Environment

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between equilibrium concentrations, albeit it is purely empirical. In Figure 11, the points that are in equilibrium with pure ternary cocrystal ABC are labeled as blue filled circles (some circles look skewed due to a slanting perspective). Peripheral to these blue points are eutectic points colored in magenta that are in equilibrium with mixtures of solid phases, including ABC+A2C (open inverted triangle), ABC+B2C (open triangle) and ABC+A (plus sign). The red dashed line stands for 1:1:1 stoichiometry line radiating from the origin. It crosses the fitting surface at a point within the region encircled by eutectic points.

Figure 11 Concentrations of isoniazid (A), nicotinamide (B) and succinic acid (C) in equilibrium with various solid phases at 40 °C. The equilibrium points in A-C plane for binary cocrystal A2C are labeled as black filled squares that are threaded together. Likewise, the equilibrium points in B-C plane for binary cocrystal B2C are labeled as black filled diamonds threaded together. The points in equilibrium with pure B2C also appear outside B-C plane as those labeled by black open diamonds. The points in equilibrium with pure ABC are labeled as blue filled circles. Surrounding the blue circles are eutectic points in equilibrium with solid mixtures containing ABC. The dashed red line stands for the 1:1:1 stoichiometry line (molar ratio) radiating from the origin. The equilibrium points for binary cocrystal systems, A2C and B2C, fall in A-C plane and B-C plane, respectively. The points in each plane are threaded for tracking. The points in A-C plane 17 ACS Paragon Plus Environment


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are labeled as black filled squares and points in B-C are labeled as black filled diamonds. Smaller squares in A-C plane represents points in equilibrium with pure A2C and larger squares do eutectic points. It is worthwhile to note that pure binary cocrystal B2C can exist outside B-C plane, as indicated by the open diamonds at the lower right corner of Figure 11. The coordinates of all points can be seen more clearly in the projections of 3D diagram on A-B, A-C and B-C planes as displayed in Figure 12 (a), (b) and (c) respectively.

(a)

(b)


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(c)

Figure 12 Projections of the 3D phase diagram 40 °C on A-B plane (a), A-C plane (b), B-C plane (c). The labels have the same denotations as in Figure 11.

Among the ten possible types of eutectic points listed in Section 2.2, only three were obtained, i.e., ABC+A, ABC+A2C and ABC+B2C. Others were not reached due to the relative stability of solid phases, or insufficient coverage of experimental conditions. For example, B crystals could coexist with neither ABC, nor a mixture of B2C and ABC, because all B crystals added could dissolve completely in a suspension of ABC, which caused the generation of B2C cocrystal and dissolution of ABC (Figure 5). In contrast, when excess A crystals were added to a suspension of ABC, only part of them dissolved and the suspension ended with a mixture of A+ABC (Figure 8). A2C cocrystal did not appear. When C crystals were added to a suspension of ABC, a different phenomenon occurred than when A or B crystals were added (Figure 6 and Figure 7). All C crystals added in the experiment dissolved. However, increase in concentration of C did not lead to generation of B2C cocrystal and the solid phase of suspension contained pure ABC. It is not clear what will happen if more C crystals are added than in Figure 6.



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It is not surprising that the type of eutectic points that are in equilibrium with three solid phases simultaneously were not obtained, such as ABC+A2C+A, ABC+A2C+C, and ABC+B2C+C. Phase rule states that F=M-P+2 Where F is the degree of freedom, M is the number of components, and P, the number of phases. The value 2 stands for temperature and pressure. In this study, temperature and pressure were fixed and M=4 (solvent, A, B and C). If three solid phases coexist in the suspension, then F=0. It means that the eutectic points of this type appear at unique locations in the 3D phase diagram and are difficult to hit during experiment. Phase rule also dictates that F is equal to 2 for suspensions with pure ABC crystals, which means that a surface exists in the phase diagram wherein pure ABC cocrystal can be obtained. Meanwhile, F=1 for suspensions with a mixture of two solid phases, which means there exist a eutectic line for every pair of solid phases such as ABC+A, ABC+A2C and ABC+B2C. Regarding the points that are in equilibrium with pure B2C outside B-C plane in the phase diagram (labeled as open diamonds), it can be seen that C concentration of these points is quite low, while the concentration of B is much higher than A concentration. These points were obtained by successively adding A2C cocrystal to a suspension of B2C solids that did not contain A in liquid phase initially. It was found that after adding a certain amount of A2C solids, ABC started to appear in the suspension and eutectic points of ABC+B2C (magenta open triangles) were reached. The stoichiometry line (red dashed line) crosses the region where pure ABC cocrystals exist, which means that evaporation of ethanol solution at 40 °C that contain equal molar of A, B and C will result in production of pure ABC, which is in agreement with the screening experiment reported earlier. With a phase diagram like the one in Figure 11, a cooling cocrystallisation process can be designed that will produce pure ternary cocrystal. In the first place, one point will be chosen in 20 ACS Paragon Plus Environment

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the region occupied by blue circles (let us assume this is at the final temperature of cooling crystallization). The location of this point should be safely away from surrounding eutectic points to avoid contamination. Then a straight line with a slope corresponding to the stoichiometry will be drawn from this point away from the origin. The other end of this line is the concentrations of A, B and C in the starting solution at a higher temperature. The length of this line depends on the solubility of ABC and the allowable maximum temperature, i.e., a clear solution should be effected at the maximum temperature when all starting solids are added. Such a stating solution thermodynamically guarantees that the final product will be pure cocrystal ABC when it is cooled to the final temperature. Other quality attributes of cocrystal ABC depend on the kinetics of cocrystallisation, which can be manipulated via techniques such as cooling rate, seeding etc. This phase diagram also suggests the necessity of using multiple solvents and multiple molar ratios of constituent components in solution-based ternary cocrystal screening such as slow evaporation. In a different solvent, the phase diagram may take a very different shape and orientation. Then it is possible that pure ternary cocrystal will not result unless the starting molar ratio of A, B and C is away from 1:1:1. 4

Conclusion

A 3-D phase diagram has been charted for isoniazid-nicotinamide-succinic acid system at 40 ˚C by using ATR-FTIR. Each axis stands for the concentration of one constituent component in this diagram. A surface region have been identified in the diagram where ternary cocrystal ABC is stable. This region is bordered by multiple eutectic points where two solid phases coexist, including ABC+A2C, ABC+B2C and ABC+A. Eutectic points that are in equilibrium with three solid phases were not reached due to their uniqueness. A phase diagram like this provides a thermodynamic basis for cocrystallisation process development. Supporting Information



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A movie is available in supporting information showing the 3D phase diagram from different angles. References 1. Schultheiss, N.; Newman, A., Pharmaceutical Cocrystals and Their Physicochemical Properties. Cryst. Growth Des. 2009, 9, 2950-2967. 2. Bolla, G.; Nangia, A., Pharmaceutical cocrystals: walking the talk. Chemical Communications 2016, 52, 8342-8360. 3. Berry, D. J.; Steed, J. W., Pharmaceutical cocrystals, salts and multicomponent systems; intermolecular interactions and property based design. Adv. Drug Deliv. Rev. 2017, 117, 3-24. 4. Swapna, B.; Maddileti, D.; Nangia, A., Cocrystals of the Tuberculosis Drug Isoniazid: Polymorphism, Isostructurality, and Stability. Cryst. Growth Des. 2014, 14, 5991-6005. 5. Childs, S. L.; Kandi, P.; Lingireddy, S. R., Formulation of a Danazol Cocrystal with Controlled Supersaturation Plays an Essential Role in Improving Bioavailability. Molecular Pharmaceutics 2013, 10, 3112-3127. 6. Drozd, K. V.; Manin, A. N.; Churakov, A. V.; Perlovich, G. L., Drug-drug cocrystals of antituberculous 4-aminosalicylic acid: Screening, crystal structures, thermochemical and solubility studies. Eur. J. Pharm. Sci. 2017, 99, 228-239. 7. Aitipamula, S.; Chow, P. S.; Tan, R. B. H., Trimorphs of a pharmaceutical cocrystal involving two active pharmaceutical ingredients: potential relevance to combination drugs. Crystengcomm 2009, 11, 1823-1827. 8. Malamatari, M.; Ross, S. A.; Douroumis, D.; Velaga, S. P., Experimental cocrystal screening and solution based scale-up cocrystallization methods. Adv. Drug Deliv. Rev. 2017, 117, 162-177. 9. Douroumis, D.; Ross, S. A.; Nokhodchi, A., Advanced methodologies for cocrystal synthesis. Adv. Drug Deliv. Rev. 2017, 117, 178-195. 10. Aitipamula, S.; Wong, A. B. H.; Chow, P. S.; Tan, R. B. H., Novel solid forms of the antituberculosis drug, Isoniazid: ternary and polymorphic cocrystals. Crystengcomm 2013, 15, 58775887. 11. Bolla, G.; Nangia, A., Binary and ternary cocrystals of sulfa drug acetazolamide with pyridine carboxamides and cyclic amides. Iucrj 2016, 3, 152-160. 12. Allu, S.; Bolla, G.; Tothadi, S.; Nangia, A., Supramolecular Synthons in Bumetanide Cocrystals and Ternary Products. Cryst. Growth Des. 2017, 17, 4225-4236. 13. Liu, F.; Song, Y.; Liu, Y. N.; Li, Y. T.; Wu, Z. Y.; Yan, C. W., Drug-Bridge-Drug Ternary Cocrystallization Strategy for Antituberculosis Drugs Combination. Cryst. Growth Des. 2018, 18, 1283-1286. 14. Tothadi, S.; Sanphui, P.; Desiraju, G. R., Obtaining Synthon Modularity in Ternary Cocrystals with Hydrogen Bonds and Halogen Bonds. Cryst. Growth Des. 2014, 14, 5293-5302. 15. Topic, F.; Rissanen, K., Systematic Construction of Ternary Cocrystals by Orthogonal and Robust Hydrogen and Halogen Bonds. Journal of the American Chemical Society 2016, 138, 6610-6616. 16. Yu, Z. Q.; Chow, P. S.; Tan, R. B. H., Operating Regions in Cooling Cocrystallization of Caffeine and Glutaric Acid in Acetonitrile. Cryst. Growth Des. 2010, 10, 2382-2387. 17. Yu, Z. Q.; Chow, P. S.; Tan, R. B. H.; Ang, W. H., Supersaturation Control in Cooling Polymorphic Co-Crystallization of Caffeine and Glutaric Acid. Cryst. Growth Des. 2011, 11, 4525-4532. 18. Yu, Z. Q.; Chow, P. S.; Tan, R. B. H., Design Space for Polymorphic Co-crystallization: Incorporating Process Model Uncertainty and Operational Variability. Cryst. Growth Des. 2014, 14, 3949-3957. 22 ACS Paragon Plus Environment

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19. Harmsen, B.; Leyssens, T., Dual-Drug Chiral Resolution: Enantiospecific Cocrystallization of (S)-Ibuprofen Using Levetiracetam. Cryst. Growth Des. 2018, 18, 441-448. 20. Chiarella, R. A.; Davey, R. J.; Peterson, M. L., Making Co-Crystals-The Utility of Ternary Phase Diagrams. Cryst. Growth Des. 2007, 7, 1223-1226. 21. Cherukuvada, S.; Nangia, A., Fast dissolving eutectic compositions of two anti-tubercular drugs. Crystengcomm 2012, 14, 2579-2588. 22. Thompson, L. J.; Voguri, R. S.; Cowell, A.; Male, L.; Tremayne, M., The cocrystal nicotinamide-succinic acid (2/1). Acta Crystallographica Section C-Crystal Structure Communications 2010, 66, o421-o424. 23. Togkalidou, T.; Fujiwara, M.; Patel, S.; Braatz, R. D., Solute concentration prediction using chemometrics and ATR-FTIR spectroscopy. Journal of Crystal Growth 2001, 231, 534543.



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For Table of Contents Use Only Elucidating the Complex Phase Behavior of a Cocrystal System Containing Two APIs and One Coformer Maria Paulene Abundo, Zai-Qun Yu, Pui Shan Chow, Reginald B. H. Tan

3D phase diagram for ternary cocrystal isoniazid(A)-nicotinamide(B)-succinic acid(C) in ethanol at 40 °C. The three axis stand for the respective concentration of A, B and C in liquid phase. A fitting surface for all points is drawn to guide the eye. The dashed red line stands for the 1:1:1 stoichiometry line (molar ratio) radiating from the origin.


Elucidating the Complex Phase Behavior of a Cocrystal System

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