The Use of Structural Data to Aid in the Identification of the Most Stable Form
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The ICH guidelines provides assistance on setting and justification of acceptance criteria and the selection of test procedures for new drug substances with the aim to generate safe drug products. The guidelines provide a series of decision trees that may help researchers to conduct the correct experiments to understand and mitigate risks. Decision tree #4 deals with polymorphs and the first box suggest performing a polymorph screen.

Figure 1. Part 1 of decision tree #4 taken from the ICH guideline Q6A.

The aim of this polymorph screen is to identify whether multiple polymorphs exist. Very often the answer to this question is “yes” and the guidelines then lead you to the next decision tree which guides you to the process of selecting the optimal crystal form of the drug substance. In many cases the thermodynamically most stable polymorph will be the desired crystalline from for development.

Selection may be achieved by applying the Burger-Ramberger rules which state that in a monotropic system the most stable form is the polymorph that has the highest melting point whilst in an enantiotropic system the polymorph with the higher heat of fusion is the more stable form (Burger, A., & Ramberger, R., 1979).

To establish the thermodynamically most stable form competitive slurry conversion experiments are often conducted in which suspensions of mixtures of two or more polymorphs are compared against solvents at different temperatures. At regular intervals samples are taken and analyzed by X-ray powder diffraction or other techniques that determine the specific solid form. This approach is not always as straightforward as it sounds and may be time consuming.

As an alternative the use of structural data may answer the question of the thermodynamically most stable and do so faster than competitive slurries. Single crystal X-ray diffraction studies provide insights of the conformation of the Active Pharmaceutical Ingredient (API) and the geometry of the intermolecular interactions, such as hydrogen bonds. Based on structural information the lattice energy can be calculated providing information regarding the stability of the form. Unfortunately, having single crystal data of multiple polymorphic forms of an API is rare but high quality X-ray powder diffraction data can easily be collected from multiple forms. If there is a possibility of obtaining the unit cell parameters it is also possible to assume the stability of the investigate forms based directly on the density of the crystals. Knowing the unit cell dimensions a, b, c and a, b, g, as well as, the molecular weight of the molecule that forms the crystal the density of the crystal can be calculated using the following equation:


d - crystal density; Fw- Formula; Weight z - number of molecules in the asymmetric unit; V- volume of the crystal; N - Avogadro constant 6.02•1023

Calculating the densities of each polymorph and applying the rule of closest packing stating that in the crystal the molecules tend to be as close as possible to achieve the lowest value of the potential energy it can be assumed that the most stable form is the polymorph with the highest density.

As an example, isonicotinamide is known to have five different forms which differ not only in the geometry of the intermolecular hydrogen bonds but also the conformation of isonicotinamide varies. In this case the most stable polymorph is Form I (d = 1.40 g/cm3), and the least stable polymorph is Form V (d = 1.34 g/cm3) The density of the other polymorphs are: Form II: d = 1.35 g/cm3, Form III: d = 1.36 g/cm3 and Form IV: d = 1. 36 g/cm3 (Aakeröy, C. B. et al, 2003; Li, J. et al, 2011; Eccles, K. S. et al, 2011).

To date four solvent free forms of carbamazepine have been identified and published. In all of these polymorphs the amide groups form a dimmer-like structure. The differences are observed in the conformation of the API and packing arrangements. The investigation of the form stability (Grzesiak, A. L. et al, 2003) revealed that the most stable polymorph is Form III (d = 1.34 g/cm3) followed by Form I (d = 1.31 g/cm3), Form IV (d = 1.29 g/cm3) and Form II (d = 1.24 g/cm3) (Figure 2).

Figure 2. Enthalpy of Melting for different Carbamazepine solvent free forms versus density (blue – Form III, red – Form I, green – Form IV, violet - Form II)

Although the approach of using the calculated density of a crystal from powder diffraction data may be attractive it also has its limitations. For instance, the calculations do not take into account the lowering of the lattice energy by a favorable hydrogen bond pattern. This may be an issue with hydrates or solvates in which the solvent molecule may facilitate a more stable hydrogen bond pattern compared to non-solvated form even when the solvate has a lower density. The model also does not include the energy profile of different conformers.

A polymorph investigation at Crystallics on aripiprazole revealed the existence of at least two polymorphs. Indexing of the X-ray powder diffraction patterns of these forms resulted in a monoclinic and triclinic crystal system. The calculated densities suggest that the monoclinic form with a density of 1.31 g/cm3 is thermodynamically more stable then the triclinic form (d = 1.29 g/cm3). Competitive slurry experiments confirmed this finding.

The use of powder diffraction data to determine the density of the crystal and through that the relative stability of a polymorph is particular useful in the early stages of development when little amount of the API is available. Determination of the thermodynamically most stable polymorph through competitive slurry experiments cost significant amounts of API. Analyzing the powder data only requires a few milligrams of material. Starting your pre-clinical phase with a stable crystalline form of your API will ensure consistent data throughout all in vitro and in vivo studies.

References: Aakeröy, C. B., Beatty, A. M., Helfrich, B. A., & Nieuwenhuyzen, M. (2003). Do polymorphic compounds make good cocrystallizing agents? A structural case study that demonstrates the importance of synthon flexibility. Crystal growth & design, 3(2), 159-165. Burger, A., & Ramberger, R. (1979). On the polymorphism of pharmaceuticals and other molecular crystals. I. Microchimica Acta, 72(3-4), 259-271. Grzesiak, A. L., Lang, M., Kim, K., & Matzger, A. J. (2003). Comparison of the four anhydrous polymorphs of carbamazepine and the crystal structure of form I. Journal of pharmaceutical sciences, 92(11), 2260-2271. Eccles, K. S., Deasy, R. E., Fábián, L., Braun, D. E., Maguire, A. R., & Lawrence, S. E. (2011). Expanding the crystal landscape of isonicotinamide: concomitant polymorphism and co-crystallisation. CrystEngComm, 13(23), 6923-6925. Li, J., Bourne, S. A., & Caira, M. R. (2011). New polymorphs of isonicotinamide and nicotinamide. Chemical Communications, 47(5), 1530-1532.