Low-Dose Scanning Electron Diffraction Microscopy of Mechanochemically Nanostructured Pharmaceuticals

Abstract

The solid-state structure of active pharmaceutical ingredients (APIs) is typically characterized in terms of polymorphism, co-crystallization, salt formation and solvate formation, which are known to have a significant effect on physical, chemical, mechanical and biopharmaceutical properties [1]. These structural features are studied in great detail, primarily using X-ray diffraction, optical spectroscopy, thermal analysis and nuclear magnetic resonance spectroscopy [1]. However, structural heterogeneity on the nanoscale (i.e. nanostructure) including crystal defects such as dislocations and stacking faults also effect properties but are challenging to study with these bulk techniques [1]. (Scanning) transmission electron microscopy ((S)TEM) is a natural choice for nanoscale characterization but, until recently, beam damage has been a major limiting factor in application to organic solids, including most APIs. Here, we report on the continued development of low-dose scanning electron diffraction (SED) microscopy, which has recently been shown to be a successful route to overcome these limitations [2, 3], and its application to reveal nanostructure in mechanochemically transformed pharmaceutical powders. SED microscopy involves the acquisition of a two-dimensional electron diffraction pattern at every probe position as a nm-sized electron probe is scanned across the specimen. Low-dose SED was performed with an electron fluence of ~ 5 e/Å 2 and a spatial resolution of ~ 5 nm by making use of a counting type Merlin/Medipix direct electron detector. A beam with ~ 0.6 mrad convergence angle was used to avoid overlapping diffraction disks and with these conditions it was typically possible to obtain a reciprocal space structural resolution of ~ 1 Å-1 , as in Figure 1a. The 4D-SED data, thus obtained, contain a wealth of spatially resolved crystallographic data, which may be analyzed in real and reciprocal space. Combined with a wealth of computational analysis, which here includes post-facto imaging and newly developed orientation mapping methods specifically for low-signal data, the technique is a highly versatile hybrid diffraction-imaging microscopy. Pharmaceutical nanostructure was explored by applying SED microscopy to mechanochemically transformed powders produced using various ball milling conditions applied to pure APIs, paracetamol:co-former mixtures [4], and mixtures in the paracetamol-theophylline-caffeine system. All pure compounds were purchased off-the shelf and the mechanical treatment was intended to induce a range of solid-state processes and reactions, which may be considered mechanochemical. This parallels the increasing use of mechanochemical methods in the pharmaceutical context to screen solid forms of APIs and their co-crystals [5] and is of fundamental interest since many industrial processing routes involve mechanical processes, e.g. milling and blending, which could potentially induce 1746