School of Chemical and Process Engineering, University of Leeds, Leeds, UK.
J Microsc. 2020 Sep;279(3):197-206. doi: 10.1111/jmi.12866. Epub 2020 Feb 18.
Formulation processing of organic crystalline compounds can have a significant effect on drug properties, such as dissolution rate or tablet strength/hardness. Transmission electron microscopy (TEM) has the potential to resolve the atomic lattice of these crystalline compounds and, for example, identify the defect density on a particular crystal face, provided that the sensitivity of these crystals to irradiation by high-energy electrons can be overcome. Here, we acquire high-resolution (HR) lattice images of the compound furosemide using two different methods: low-dose HRTEM and bright-field (BF) scanning TEM (STEM) scanning moiré fringes (SMFs). Before acquiring HRTEM images of furosemide, a model system of crocidolite (asbestos) was used to determine the electron flux/fluence limits of low-dose HR imaging for our scintillator-based, complementary metal-oxide semiconductor (CMOS) electron camera by testing a variety of electron flux and total electron fluence regimes. An electron flux of 10 e /(Å s) and total fluence of 10 e /Å was shown to provide sufficient contrast and signal-to-noise ratio to resolve 0.30 nm lattice spacings in crocidolite at 300 kV. These parameters were then used to image furosemide which has a critical electron fluence for damage of ≥10 e /Å at 300 kV. The resulting HRTEM image of a furosemide crystal shows only a small portion of the total crystal exhibiting lattice fringes, likely due to irradiation damage during acquisition close to the compound's critical fluence. BF-STEM SMF images of furosemide were acquired at a lower electron fluence (1.8 e /Å ), while still indirectly resolving HR details of the (001) lattice. Several different SMFs were observed with minor variations in the size and angle, suggesting strain due to defects within the crystal. Overall BF-STEM SMFs appear to be more useful than BF-STEM or HRTEM (with a CMOS camera) for imaging the crystal lattice of very beam-sensitive materials since a lower electron fluence is required to reveal the lattice. BF-STEM SMFs may thus prove useful in improving the understanding of crystallization pathways in organic compounds, degradation in pharmaceutical formulations and the effect of defects on the dissolution rate of different crystal faces. Further work is, however, required to quantitatively determine properties such as the defect density or the amount of relative strain from a BF-STEM SMF image.
有机晶体化合物的制剂加工对药物性质有重大影响，例如溶解速率或片剂强度/硬度。透射电子显微镜（TEM）有潜力解析这些晶体化合物的原子晶格，例如，确定特定晶面的缺陷密度，前提是这些晶体对高能电子的辐照的敏感性可以克服。在这里，我们使用两种不同的方法获取呋塞米的高分辨率（HR）晶格图像：低剂量 HRTEM 和明场（BF）扫描透射电子显微镜（STEM）扫描莫尔条纹（SMF）。在获取呋塞米的 HRTEM 图像之前，我们使用青石棉（石棉）模型系统来确定低剂量 HR 成像的电子通量/通量限制对于我们基于闪烁体的互补金属氧化物半导体（CMOS）电子相机，通过测试各种电子通量和总电子通量范围。结果表明，在 300kV 时，电子通量为 10e /（Å s）且总通量为 10e /Å 足以提供足够的对比度和信噪比，以分辨青石棉中 0.30nm 的晶格间距。然后使用这些参数对呋塞米进行成像，在 300kV 时，其临界电子通量损伤大于 10e /Å 。呋塞米晶体的 HRTEM 图像仅显示了总晶体的一小部分显示晶格条纹，这可能是由于在接近化合物临界通量的采集过程中受到辐照损伤。在较低的电子通量（1.8e /Å ）下获取了呋塞米的 BF-STEM SMF 图像，同时仍间接解析了（001）晶格的 HR 细节。观察到几个不同的 SMF，大小和角度略有变化，这表明晶体内部存在缺陷导致应变。总体而言，与 BF-STEM 或 HRTEM（使用 CMOS 相机）相比，BF-STEM SMF 更适合对非常敏感的光束材料的晶格成像，因为需要较低的电子通量才能显示晶格。因此，BF-STEM SMF 可能有助于提高对有机化合物结晶途径、药物制剂降解以及不同晶面溶解速率的缺陷的理解。然而，需要进一步的工作来从 BF-STEM SMF 图像定量确定诸如缺陷密度或相对应变量等特性。
