Design and synthesis of a library of pyrazolo[1,5-a]pyridine fluorophores for developing new fluorescent probes

Fluorescence imaging is a non-invasive technique useful to understand and visualize biological processes in living organisms.1 Most of the available dyes normally used for this aim, suffer for various limitations such as large size, poor water solubility or low cell permeability.2 In recent years, several research has been focused on developing novel small fluorescent dyes with low molecular weight, good signal to noise ratio and optimized proprieties that can be applied to most biological samples without disturbing the physiological envirorment.3 Pyrazolo[1,5-a]pyridin-2-ol is a small scaffold with a strong emission in the blue region, recently studied as bioisoster of the carboxylic acid moiety.4,5 With the purpose to increase the fluorescent properties of this scaffold, different strategies were investigating. Generally, the donor–π-acceptor moiety is the key structural framework modulated to give fluorescence to small organic compounds. To improve the intrinsic properties of our probe (absorption/emission/Φ/Stokes shift), the latter was modulated with different stronger electronic drawing and accepting groups. Furthermore, the scaffold was studied by expanding the π-conjugated network with the introduction of a wider structure to improve the emission profile of the system (see Figure). Preliminary data of the fluorescent proprieties of the designed target compounds, showed a bathochromic (red) shift in the UV–vis absorption and emission spectra. Further studies need to be conducted to establish the best attachment position to use our molecule as an imaging probe. Theoretical design, synthesis and fluorescent proprieties will be here presented and discussed.