Surface-assisted laser desorption/ionization mass spectrometry (SALDI-MS) emerges as an effective method for analyzing small molecules. At the current stage, its application is still limited by its detection sensitivity. According to the mechanism of SALDI-MS, the sensitivity highly depends on the laser desorption/ionization (LDI) efficiency. In this study, we adjust the localized surface plasmon resonance (LSPR) between particles by changing the gap, thereby improving the efficiency of LDI. The gaps of nanoparticles can be adjusted by the ionic strength in the solution. With the decrease of the gaps, the thermal effect of the nanoparticles is gradually enhanced and the number of excited electrons is also increased, which improves the sensitivity of SALDI-MS. The prepared substrate with optimized gaps has wide applicability for the detection of small biomolecules and dye molecules. Moreover, the substrate can successfully detect trace small molecules in complex samples. The analysis of sulfonamide in lake water shows the commendable performance of the substrate. The relative standard deviations (RSD) of spot-to-spot and batch-to-batch are 3.73% and 7.22% respectively, confirming the high reproducibility and the detection limit is as low as 100 pM, indicating the high sensitivity of the substrate. The results demonstrate that adjusting the LSPR of metal nanoparticles is a promising method for improving the performance of SALDI-MS.