A simple "OFF-ON" fluorescent system was proposed for selective and sensitive detection of ferric ion (Fe) and pyrophosphate (PPi) in living cells. The method was constructed based on the bright yellow emission of carbon dots (y-CDs), which were prepared using o-phenylenediamine (OPD) as the precursor via a facile hydrothermal treatment. The as-obtained y-CDs, with an average size of 2.6 nm, exhibited an excitation-independent emission peak at 574 nm. The fluorescence of y-CDs can be remarkably quenched by Fe with high selectivity and sensitivity. Interestingly, the quenched fluorescence can be recovered regularly upon addition of PPi, showing a promising detection for PPi. The linear ranges for Fe and PPi detections were 0.05-80 and 0.5-120 μM, respectively, and the corresponding limit of detections (LODs) were 22.1 and 73.9 nM. As we proved the y-CDs have negligible cytotoxicity and excellent biocompatibility, further application to the fluorescence imaging of intracellular Fe and PPi were conducted, suggesting the prepared y-CDs can be used to monitor Fe and PPi variation in living cells. Overall, our developed y-CDs-based OFF-ON switch fluorescent probe has the advantages of simplicity, agility, high sensitivity and selectivity, which provides a promising platform for environmental and biology applications, and paves a new avenue for monitoring the hydrolysis process of adenosine triphosphate disodium salt (ATP) by detection of PPi in organisms.