Chemically Specific Coherent Raman Imaging of Liquid–LiquidPhase Separation and Its Sequelae

Liquid–liquid phase separation (LLPS) plays a pivotal role in processes ranging from cellular structure reorganization to the formation of crystalline structures in materials science. In the pharmaceutical field, it has been demonstrated to impact drug crystallization and delivery. To date, characterization of LLPS has been limited to nonspatially resolved or nonchemically resolved analyses. In this study, we employed chemically specific stimulated Raman scattering (SRS), combined with second harmonic generation (SHG), for the first time to image crystallization in the presence of LLPS. Using the model compound ibuprofen, we examined the interplay between LLPS and crystallization, and explored the influence of both dissolution medium and enantiomeric form on this behavior. In doing so, we also discovered and partially characterized a new polymorph of (S)-ibuprofen. Our results demonstrate the potential of correlative SRS and SHG for monitoring phase separation and crystallization in real time, giving mechanistic insights into the spatial distribution, chemical composition and structure of phase-separated domains and newly formed crystals. Such insights could benefit not only pharmaceutical development, but also the biomedical, food and chemical sectors.