We report the development and testing of an accurate mass–time tag library for plant natural product profiling. Display omitted ► The development of an accurate mass–time tag approach for the identification of plant natural products is reported. ► LC/MS and MS/MS data sets were integrated into online spectral search tools and repositories. ► The utility was demonstrated by the detection of active principles in 27 medicinal plant species. We report the development and testing of an accurate mass–time (AMT) tag approach for the LC/MS-based identification of plant natural products (PNPs) in complex extracts. An AMT tag library was developed for approximately 500 PNPs with diverse chemical structures, detected in electrospray and atmospheric pressure chemical ionization modes (both positive and negative polarities). In addition, to enable peak annotations with high confidence, MS/MS spectra were acquired with three different fragmentation energies. The LC/MS and MS/MS data sets were integrated into online spectral search tools and repositories (Spektraris and MassBank), thus allowing users to interrogate their own data sets for the potential presence of PNPs. The utility of the AMT tag library approach is demonstrated by the detection and annotation of active principles in 27 different medicinal plant species with diverse chemical constituents.