Database Enables UHPLC-Q-Orbitrap-MS/MS Method to Discover 16 "Overlooked" Compounds and Supply Three New Pharmacopoeia Quality-markers for Charred Semen Arecae (Jiaobinglang)

Abstract

Background The emerging UHPLC-Q-Orbitrap-MS/MS method is not good at deep analysis of charred Chinese herbal medicines (CHMs), for the absence of database. The study tried to create a new strategy, i.e., database-affinity ultra-high-performance liquid chromatography-quadrupole-orbitrap-tandem mass spectrometry (UHPLC-Q-Orbitrap-MS/MS), to deeply analyze a CHM instance, i.e., charred Semen Arecae (CSA). Methods The database-affinity UHPLC-Q-Orbitrap-MS/MS was used to qualitatively and quantitatively analyze the lyophilized aqueous extract of CSA. Relative chemical contents and antioxidant activities of compounds were further evaluated via PTIO• radical-scavenging assay. Antioxidant contributions were calculated as "relative antioxidant level × chemical content." Based on chemical and antioxidant analyses, supplemental Q-markers were proposed and thereafter investigated for their traceability using quantum chemistry. Furthermore, they were validated for the applicability using multiple products and multiple analytical technologies (LC-MS and HPLC-UV). Results Thirty-five compounds were putatively identified by database-affinity UHPLC-Q-Orbitrap-MS/MS; especially, 16 "Overlooked, " compounds were discovered in CSA. Three procyanidin isomers (B1, B2, B3) were differentiated from each other and the Maillard reaction intermediate 5-hydroxymethylfurfural (5-HMF) was found to enrich in CSA. (+)-Catechin and (-)-epicatechin exhibited the highest antioxidant contributions. (+)-Catechin, (-)-epicatechin, and 5-HMF were thus proposed as CSA’s supplemental Q-markers. They were evidenced to show good traceability, pharmacology-relevance, TCM relevance, testability, and specificity. It was also demonstrated to be applicable for various CSA products from different batches (in total 25 batches), different locations, different extraction methods, and even different analytical technologies. Conclusion Sixteen "Overlooked, " compounds are found in CSA for the first time by database-affinity UHPLC-Q-Orbitrap-MS/MS. Three supplemental Q-markers should be integrated with arecoline, to construct a new Pharmacopeia Q-markers system. The system can overcome the limitations of the old and risky Q-marker (arecoline). This successful instance indicates that database can enable UHPLC-Q-Orbitrap-MS/MS for deep analysis of Chinese herbal medicine and quality-control update of charred CHM.