The invention discloses a fluorescent probe synthesized by rhodamine B, triethylene tetramine and phenyl isothiocyanate and a preparing method and application thereof. Specifically, firstly, the rhodamine B reacts with the triethylene tetramine to form a spirocyclic amide structure which is stable in property, nitrogenous bridged segments are introduced in, and the rhodamine B reacts with the phenyl isothiocyanate, so that sulphur atoms are introduced in. By means of the thiophil property of mercury ions, each molecule in an obtained novel rhodamine B-phenylthiourea derivative RTTU can be combined with three mercury ions, and therefore high selectivity and detection sensitivity are achieved. Fluorescent probe molecules can conduct detection in a neutral buffer solution, especially an aqueous phase and can also detect the mercury ions from mercuric chloride, and higher practicability is achieved. In addition, the fluorescent probe molecules can be directly precipitated out in the preparing process, further separation and purification are not needed, and the synthesizing method is simple and convenient and easy to implement.