Despite the key roles of L-glutathiones (GSHs) inbiology and nano-biotechnology, understanding their labile structures and hydrogen bond interactions with nanoparticles has posed a critical challenge to the scientific community. The structural conformation of GSH as a capping layer on gold nanoparticle (AuNP) and silver nanoparticle (AgNP) surfaces is investigated. In this report, we attempt to explore the material-dependent interaction of GSH with different spherical nanoparticle surfaces by employing Fourier transform infrared (FTIR) spectroscopy. The infrared signal of amide I of GSH is studied as a function of different materials' spherical nanoparticles with comparable size. We revealed the β-sheet secondary structure of GSH on AgNPs and the random structure on AuNPs even though both the nanoparticles have comparable shapes and sizes and belong to the same group of the periodic table. The GSH is firmly anchored on the gold and silver surface via the thiol of the cys part. However, our experimental data designate a further interaction with the AgNP surface via the carboxylic acid group of the gly-and glu-end of the molecule. It is observed that enhancement of IR absorption of amide I of GSH is pronounced by a factor of 10 on AuNP but, in contrast, on the same-sized AgNP, the suppression is perceived by a factor of 2, even though both are plasmonic materials with respect to free GSH. This study can be used as a point of reference for understanding the structural conformation of the capping layer on nanoparticle surfaces as well as surface enhancement of the IR absorption of amide I. We would like to emphasize that molecular self-assembly on the nanoparticle surfaces is definitely of very broad interest for chemists working in nearly any subdiscipline, spanning from the nanoparticle-based medicine to surfaceenhanced spectroscopy to heterogeneous catalysis, etc.