Utilizing metabolomics and network analysis to explore the effects of artificial production methods on the chemical composition and activity of agarwood

Introduction: Agarwood is a traditional aromatic southern medicine. It has a long history of being used in traditional Chinese aromatherapy to treat insomnia, anxiety and depression. Due to the scarcity of wild resources, people have planted trees successfully and begun to explore various agarwood-inducing techniques. This study comparative analysis of volatile metabolites in agarwood produced by various inducing techniques and its potential sleep-promoting, anti-anxiety and anti-depressant network pharmacological activities. Methods: A total of 23 batches of two types of agarwood were collected, one of which was produced by artificial techniques, including 6 batches of TongTi (TT) agarwood produced by “Agar-Wit” and 6 batches of HuoLao (HL) agarwood produced by “burning, chisel and drilling”, while the other was collected from the wild, including 6 batches of BanTou (BT) agarwood with trunks broken due to natural or man-made factors and 5 batches of ChongLou (CL) agarwood with trunks damaged by moth worms. The study employed metabolomics combined with network analysis to compare the differences in volatile metabolites of agarwood produced by four commonly used inducing techniques, and explored their potential roles and possible action targets in promoting sleep, reducing anxiety, and alleviating depression. Results: A total of 147 volatile metabolites were detected in agarwood samples, mainly including small aromatic hydrocarbons, sesquiterpenes and 2-(2-phenylethyl) chromone and their pyrolysis products. The results showed composition of metabolites was minimally influenced by the agarwood induction method. However, their concentrations exhibited significant variations, with 17 metabolites showing major differences. The two most distinct metabolites were 6-methoxy-2-(2-phenylethyl) chromone and 6,7-dimethoxy-2-(2-phenylethyl) chromone. Among the volatile metabolites, 142 showed promising potential in treating insomnia, anxiety, and depression, implicating various biological and signaling pathways, predominantly ALB and TNF targets. The top three active metabolites identified were 2-(2-phenylethyl) chromone, 1,5-diphenylpent-1-en-3-one, and 6-methoxy-2-[2-(4'-methoxyphenyl) ethyl] chromone, with their relative content in the four types of agarwood being TT>HL>CL>BT. Conclusion: The differences in the content of 2-(2-phenylethyl) chromones suggest that they may be responsible for the varying therapeutic activities observed in different types of agarwood aromatherapy. This study offers theoretical support for the selection of agarwood in aromatherapy practices.