The chemical composition of herbal medicines is very complex, and their therapeutic effects are determined by multi-components with sophisticated synergistic and/or suppressive actions. Therefore, quality control of herbal medicines has been a formidable challenge. In this work, we describe a fast analytical method that can be used for quality assessment of herbal medicines. The method is based on ligand fishing using human-serum-albumin-functionalized magnetic nanoparticles (HSA-MNPs) and mass spectrometry. To demonstrate the applicability of the proposed method, eight samples of Dioscorea panthaica were analyzed. The sampled plants were of both wild and cultivated origins. They grew at different geographical locations and were harvested at different times. The ligands bound to HSA-MNPs were isolated from the plant extracts and detected by using direct infusion electrospray ionization mass spectrometry (DI-ESI-MS). Chemical identity has been confirmed for five of the ligands isolated. From more than 15 peaks in the ESI-MS spectrum, 11 common peaks were selected for calculating the correlation coefficient and cosine ratio. The values of correlation coefficient and cosine ratio were >0.9824 and >0.9988, respectively, for all the samples tested. The results indicated a high level of similarity among the eight D. panthaica samples. Compared with chromatographic fingerprint analysis, the proposed HSA-MNP-based DI-ESI-MS/MS approach was not only fast and easy to carry out but also biological-activity-oriented, promising a more effective data interpretation and thus reliable assessment conclusions.