Analyzing noncovalent interactions between notoginseng saponins and lysozyme by deposition scanning intensity fading MALDI‐TOF mass spectrometry

Abstract Analysis of noncovalent interactions between natural products and proteins is important for rapid screening of active ingredients and understanding their pharmacological activities. In this work, the intensity fading MALDI‐TOF mass spectrometry (IF‐MALDI‐MS) method with improved reproducibility was implemented to investigate the binding interactions between saponins from Panax notoginseng and lysozyme. The benchmark IF‐MALDI‐MS experiment was established using N , N ′, N ″‐triacetylchitotriose‐lysozyme as a model system. The reproducibility of ion intensities in IF‐MALDI‐MS was improved by scanning the whole sample deposition with a focused laser beam. The relative standard deviation (RSD) of deposition scanning IF‐MALDI‐MS is 5.7%. Similar decay trends of the relative intensities of notoginseng saponins against increasing amounts of lysozyme were observed for all six notoginseng saponins. The half‐maximal fading concentration (FC 50 ) was calculated to quantitatively characterize the binding affinity of each ligand based on the decay curve. According to the FC 50 values obtained, the binding affinities of the six notoginseng saponins were evaluated in the following order: notoginsenoside S > notoginsenoside Fc > ginsenoside Rb1 > ginsenoside Rd > notoginsenoside Ft1 > ginsenoside Rg1. The binding order was in accordance with molecular docking studies, which showed hydrogen bonding might play a key role in stabilizing the binding interaction. Our results demonstrated that deposition scanning IF‐MALDI‐MS can provide valuable information on the noncovalent interactions between ligands and proteins.