The invention discloses a fluorescent probe for detecting CO (carbon monoxide) in cells and a preparation method and application of the fluorescent probe. The fluorescent probe for detecting the CO in the cells is shown as the structural formula below, wherein R refers to H, -CH3, -CH2CH3 or -COOEt. According to the invention, a novel cyclopalladation radical group shown in the description below is synthesized and capable of responding to the CO more rapidly and sensitively, and after the novel cyclopalladation radical group is combined with BODIPY, selectivity for the CO is improved and the CO can be detected more rapidly and sensitively. The preparation method includes enabling paranitrobenzoyl chloride, dimethyl pyrrole and boron trifluoride diethyl etherate or the paranitrobenzoyl chloride, dimethyl pyrrole derivatives and the boron trifluoride diethyl etherate to react to generate nitrophenyl BODIPY, reducing nitro in the nitrophenyl BODIPY to amino, adding nitrite and 3,5-dimethylphenol to undergo diazotization, and adding palladium salt to cyclopalladation so as to obtain a target product, namely, the fluorescent probe for detecting the CO in the cells (ACP-CO). The preparation method is simple in synthesis method and applicable to industrial production.