Laser desorption ionization mass spectrometry (LDI-MS) is a primary tool for biological analysis. Its success relies on the use of chemical matrices that facilitate soft desorption and ionization of the biomolecules, which, however, also limits its application for metabolomics study due to the chemical interference by the matrix compounds. The requirement for sample pretreatment is also undesirable for direct sampling analysis or tissue imaging. In this study, antireflection (AR) metal surfaces were investigated as sample substrates for matrix-free LDI-MS. They were prepared through ultrafast laser processing, with high light-to-heat energy conversion efficiency. The morphology and micro/nanostructures on the metal surfaces could be adjusted and optimized by tuning the laser fabrication process. The super-high UV absorption at 97% enabled highly efficient thermal desorption and ionization of analytes. The analytical performance for the matrix-free LDI was explored by analyzing a variety of biological compounds, including carbohydrates, drugs, metabolites, and amino acids. Its applicability for direct analysis of complex biological samples was also demonstrated by direct analysis of metabolites in yeast cells.