Figure 2.

Intracellular self-assembled crystal organization of CFZ-H⁺Cl⁻. (a) CFZ-H⁺Cl⁻ chemical structure with color coding corresponding to the regions of the structure in the crystal arrangement. (b) Freeze-fracture electron micrograph of CLDIs within a Kupffer cell, showing that the CLDIs are sequestered inside the cell as multiple layered planes with lamellar spacing of 6 nm to 14 nm, by a double membrane of biological origin (~20 nm in thickness)²¹. (c) Crystal packing of CFZ-H⁺Cl⁻ showing the three crystallographic planes of the crystal: a hydrophobic face in orange (plane 1), a hydrophilic face in blue (plane 2), and an amphipathic face (plane 3)⁸. (d) CLDIs (100–1.5X magnification) zoomed in (zoom factor of 2) in alveolar macrophages show high dichroism and an axis of highest transmittance, with a different profile (represented by the different signal intensity associated with the color bar) observed specifically around the edges of the crystal. (e) Raman spectra of free base CFZ, CFZ-H⁺Cl⁻, untreated and treated alveolar macrophages (4 and 8wks CFZ-fed mice) show distinct Raman peaks distinguishing between the different forms of the drug (red in the brightfield images): free base CFZ (1341 and 1465 cm⁻¹), CFZ-H⁺Cl⁻ (1399 cm⁻¹). (f) Reflected brightfield and Raman images of alveolar macrophages from CFZ-fed mice prepared on silicon chip substrates; basis spectral fitting was used to show the temporal accumulation and cytoplasmic packaging of CFZ into highly ordered CLDIs.