Fresh ginkgo leaves were used as carbon source to synthesize carbon dots (CDs) by hydrothermal method with good water solubility and high stability. The structure and optical properties of CDs was characterized by TEM, XRD, FTIR, UV-Vis and fluorescence spectra. It was found that rutin could reduce the fluorescence intensity of CDs. Based on the quenching phenomenon, a fluorescence probe with high selectivity and sensitivity was obtained. The results showed that the quenching degree of CDs fluorescence presented a good linear relationship with rutin concentration in the range from 10 to 80 μmol/L. The equation was ln(F/F0)= -0.0175c(μmol/L)-0.01492, and the correlation coefficient R was 0.9997. The detection limit is 1.5μmol/L. Moreover, citric acid CDs was synthesized using citric acid as carbon source. These two fluorescent probes were applied to the determination of Rutin in sophora flower bud, and a close result is obtained.