Abstract: Antibiotic misuse is becoming a worldwide public health issue due to serious risk to human health and environment. Residual antibiotics in environment mainly come from the wastewater from pharmaceutical factories, aquaculture, animal husbandry and hospitals. Antibiotics are included in the newly published List of Newly Controlled Pollutants (2022 Edition) (Draft for Comments). However, test standards are still lacking. Highly accurate results of the trace amounts of antibiotics distributed in the environment can be obtained by mass spectrometry coupled with chromatography. Nonetheless, sample pretreatment is time-consuming and expert-required. Compared with laboratory detection, ambient miniature mass spectrometry can realize on-site detection of the antibiotics without or with little pretreatment. Onsite detection can be used for rapid qualitative and semi-quantitative detection, which can reflect the levels of water body pollution and accurately trace the pollutant sources. On-site detection also can provide technical support for the regulatory governance of antibiotics. But now no-studies have been reported on the on-site detection of environmental antibiotics in the field. It is urgent to develop a rapid detection method for antibiotic residues in the environment to assess antibiotic pollution and provide support for regulation. In this paper, the detection of antibiotics was carried out based on flash desorption and acetone assisted photoionization source miniature ion trap mass spectrometry, the detection parameters of antibiotics were systematically optimized including gas flow and injection time sequence, and a rapid and high throughput detection method of antibiotics was established. The efficiency of thermal desorption was improved using pulse purging mode and halogen lamp flash thermal desorption, and the signal intensities of high boiling point antibiotics were significantly increased. Quantitative detection of antibiotics was realized by isotope internal standard, and preliminary detection of antibiotics in water was carried out. This method does not need complicated pre-processing and can be completed in 1 min. The detection limits of isoniazid (INZ), ethionamide (ETN) and voriconazole (VCZ) were 636, 160, 35 μg/L, respectively, calculated based on S/N=3 at the injection volume of 1 μL. It is applicable to the field, highthroughput and highly sensitive detection of antibiotics.