Abstract: The development and application of ambient ionization and miniature mass spectrometry have become the current research hotspot, capable of rapid, sensitive, on-site, and real-time analysis of samples without complicated sample pretreatment. In this study, a rapid, on-site method was developed for the screening of hazardous substances in infant toiletries using ambient ionization and miniature mass spectrometry. Without time-consuming sample pretreatment, the liquid contained in baby wipes was drawn and then dropped onto a Grade 3MM triangle paper substrate. A metal mini-alligator clip was used to hold the triangular piece of paper with its tip about 1 cm in front of the inlet of a miniature mass spectrometer. Aliquots of 10 μL methanol containing 0.3% formic acid was used as spray solvent and dropped onto the paper substrate. A spray voltage of 3.0 kV was applied, and then charged droplets were emitted from the paper tip, leading to the acquisition of the MS/MS spectra in positive ion mode. Based on the observed signal-to-noise ratio (S/N) of the most intense MS/MS fragment peaks and using S/N=3 as a criterion, the limits of detection (LODs) of 5-chloro-2-methyl-4-isothiazolin-3-one and 2-methyl-4-isothiazolin-3-one were estimated to be 50 and 20 μg/kg, respectively. Other infant toiletry samples, including toothpaste, baby bottle and dish liquid, and nappy cream, were analyzed by extraction nano-electrospray ionization (nanoESI). A borosilicate glass capillary (1.5 mm×0.86 mm×5 cm) with a pulled tip was prepared using a P-1000 micropipette puller. The samples spiked with the analytes, such as toothpaste spiked with diethylene glycol and salicylic acid, baby bottle and dish liquid spiked with atranol and lyral, and nappy cream spiked with propylparaben and butylparaben, were gently picked up using a metal wire probe, and then inserted into the pulled capillary, which had been prefilled with 10 μL of spray solvents. The capillary was placed in front of the inlet of the miniature mass spectrometer. A high voltage of 3.0 kV in positive ion mode or -2.5 kV in negative ion mode was applied via the metal wire, producing the spray for immediate nanoESI analysis. The LODs were estimated to be between 10 and 50 μg/kg for the eight hazardous substances. Coupling with miniature mass spectrometry, the entire analysis could be completed in less than 1 min. The simplified analytical procedures effectively solve the disadvantages of traditional analytical methods that are cumbersome and time-consuming. The proposed method is simple, rapid, sensitive, efficient, and suitable for rapid, on-site screening of hazardous substances in infant toiletries.