We have applied surface-assisted laser desorption/ionization mass spectrometry (SALDI-MS) usingtitanium dioxide nanoparticles(TiO₂NPs)as the matrix and captopril (CAP) as internal standard for the determination of the concentrations of theanine and four catechins—catechin, (_)-epigallocatechin (EGC), (_)-epicatechin gallate (ECG), and (_)-epigallocatechin gallate (EGCG).Under the optimal conditions (240 nM TiO₂ NPs and 10 μM CAP), this SALDI-MS approach provides linearity of 0.3– 80 (r= 0.990), 1.2–100 (r= 0.987), 4–120 (r= 0.995), 6–120 (r= 0.983), and 2–120 μM (r= 0.991) for theanine, catechin, EGC, ECG, and EGCG, respectively. The limits of detection (LODs; S/N = 3)for theanine, catechin, EGC, ECG, and EGCG provided by this SALDI-MS approach are 0.1, 0.35, 1.0, 1.45, and 0.5 μM, respectively. This approach provides spot-to-spot and batch-to-batch variations of less than 10% and 13%, respectively, for the analysis of tea samples.With advantages of simplicity, accuracy, precision, and great reproducibility, we have applied the SALDI-MS approach for the analysis of tea samples, with identified peaks for theanine, catechin, EGC, ECG, and EGCG. Tea samples from Taiwan and four other areas have various SALDI-MS profiles, showing their potential for differentiation of tea samples from different sources. Our result also shows that tea samples harvested in different seasons and counties in Taiwan provide significantly different MS profiles. The amounts of theanine and EGC in the Oolong tea from Lishan are much higher than those in the other tea samples.