In mass spectrometry, the type and design of ionization source play a key role on the performance of a given instrument. Therefore, it is of paramount importance to evaluate newly developed sources for their suitability to analyze food contaminants like pesticide residues. Here, we carried out a head-to-head comparison of key extraction and analytical performance parameters of an electrospray ionization (ESI) source with a new atmospheric pressure ionization source, UniSpray (US). The two interfaces were evaluated in three matrices of different properties (coffee, apple, and water) to determine if multiresidue analysis of 81 pesticides by QuEChERS extraction and LC-MS/MS analysis could be improved. Depending on the matrix and irrespective of the chemical class, US provided a tremendous gain in signal intensity (22- to 32-fold in peak area, 6- to 7-fold in peak height), a threefold to fourfold increase in signal-to-noise ratio, a mild gain in the range of compounds that can be quantified, and up to twofold improvement of recovery. UniSpray offered comparable linearity and precision of the analyses with ESI, and did not affect the ion ratio. A gain in sensitivity of many compounds was observed with US, but in general, the two ionization interfaces did not show significant difference in LOD and LOQ. UniSpray suffered less signal suppression; the matrix effect was in average 3 to 4 times more pronounced, but showed better values than ESI. With no effect on recovery efficiency, US improved the overall process efficiency 3 to 4 times more than ESI. Graphical abstract.