Carbon dots (CDs) have been a new class of fascinating carbon-based fluorescent nanomaterials. In the present work, new N-doped CDs with highly crystalline graphite structures are prepared from renewable precursors, chitosan and tartaric acid, and are well characterized. The prepared CDs are applied as a biocompatible fluorescent sensor for the sequential detection of Fe3+ and AA. Among various transition metal ions, Fe3+ can selectively quench the fluorescence of CDs. Upon the further addition of AA, the quenched fluorescence of CDs is then restored as Fe3+ is reduced to Fe2+ by AA, which can be utilized for the fluorescent determination of AA. A good linear relationship in the range of 0-150 μM of AA concentration is established with a low detection limit of 0.02 µM. Moreover, the practical applications of this fluorescent sensing method in measurement of AA in food samples are successfully realized with satisfactory results.