The first benzothiazole-ratio probe, 4-(benzothiazol-2-yl)-2-hydroxy benzaldehyde (HBTA), dynamically tracks the concentration of malononitrile by showing its sensitivity to the two phases (organic and aqueous) of malonitrile. In order to explore the sensing mechanisms of probe HBTA, the density functional theory (DFT) and time-dependent DFT were used. We first demonstrated that the single fluorescence observed in the experiment was due to HBTA-Enol rather than HBTA-Keto, and observed the local excited state and charge transfer state during the excitation process of HBTA. Additionally, the sensing mechanism of HBTA in two different solvents (organic and aqueous) was studied, and we found that malononitrile could be detected by HBTA efficiently and sensitively in the physiological environment. Meanwhile, the geometric parameters and reduction density gradient were calculated. Furthermore, we used frontier molecular orbitals (FMOs) and hole-electrons to analyze the charge distribution, providing strong evidence for the possibility of excited-state intramolecular proton transfer (ESIPT) and intramolecular charge-transfer (ICT) processes occurring.