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The intracellular environment is crowded with diverse biomacrolecules (~80-400 mg/ml), likely affecting various biological processes such as protein folding, binding of small molecules, enzymatic activity, and pathological protein aggregation. As a model we have been using solutions of Ficoll, a highly branched polysaccharide, to mimic the environment. Besides its biomedical applications (e.g. blood separation), it has been used as a macromolecular crowder in studies of protein folding and stability, cell volume signaling, tissue engineering, and nanotransport. In this study, our goal is to identify and assess Raman spectral signatures associated with Ficoll molecules and Ficoll-Ficoll interactions for future investigations of crowding effects. In addition to the Raman peaks of water (~1640 cm-1 and ~3200 cm-1) and dissolved O2 (~1556 cm-1) and N2 (~2331 cm-1) we identified a distinct Raman peak (~2900 cm-1) in the 1500-3500 cm-1 wavenumber range, which is associated with Ficoll and CH and CH2 stretching modes. As the Ficoll concentration increases, the intensity of the Ficoll Raman peaks increases while the intensity of the water Raman peaks decreases, the latter likely due to reduction of water content. Further, we have applied the intensity correlation analysis (ICA) method to assess systematic changes of Raman spectra with Ficoll concentration (up to 1000 mg/ml). ICA indicates an overall linear trend over the full wavenumber range, but also shows closed loops that can be attributed to slight changes of the profiles of certain peaks. The results demonstrate ICA as a potential insightful tool for identifying Ficoll in chemical analysis of crowded biological samples.
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