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Mapping of total fluorescence of blood plasma is extensively used for its characterization and allows, with varied success, to detect pathological processes in organism (e.g., cancer). This approach is based on evaluation of blood plasma native fluorophores (proteins, NADP, flavins etc.) intensities, that depend on metabolism, and is mostly empirical. Here, we studied fluorescence spectra of blood plasma of rats with steroid-induced diabetes to determine fluorescence markers of hyperglycemia. To interpret the obtained results, model experiments with blood plasma proteins glycation were performed in vitro. We used laser fluorescence spectroscopy to monitor glucose binding sites and conformational changes of serum albumin during its incubation with glucose, including aggregates formation. In the latter case, the role of π-stacking in fluorescence response of proteins cross-linkings was studied.