Hydrogen-Bond-Assisted Dual-Mode Ratiometric Detection of Uric Acid Using Pyrimidine-Driven Charge Transfer Probes: From Mechanistic Studies to Real-Life Applications

The study highlights the design, synthesis, and photophysical evaluation of two pyrimidine-based charge transfer probes (compounds <b>1</b> and <b>2</b>) for the selective detection of uric acid (UA) in aqueous environments and biological fluids. Both probes exhibited distinct spectral responses attributable to their structural differences, with compound <b>1</b> demonstrating superior sensitivity. Quantitative analysis revealed a strong linear correlation (<i>R</i>² = 0.998) between UA concentration and fluorescence intensity, enabling detection as low as 0.12 μM, which is well within the clinically relevant range. Mechanistic investigations confirmed the indispensable role of the pyrimidine unit in forming multipoint hydrogen-bonding interactions with UA, which induced spectroscopic changes and disrupted probe aggregates. Environmental factors such as pH, polarity, and temperature were systematically examined, revealing that probe performance was optimal at moderate pH (4-8) and 60-70% water content, with elevated temperatures (50 °C) enhancing the response due to the dissociation of supramolecular aggregates. Screening in diluted human urine validated the practical applicability of the probe, demonstrating high recovery rates and minimal matrix interference. The compound was also utilized for the detection of UA in HeLa cells. Also, chemically modified paper strips were developed for rapid, on-site detection of UA.