Revealing of Intracellular Antioxidants in Dendrobium nobile by High Performance Liquid Chromatography-Tandem Mass Spectrometry

Abstract

The medicinal plant Dendrobium nobile is an important natural antioxidant resource. To reveal the antioxidants of D. nobile, high performance liquid chromatography-tandem mass spectrometry (HPLC-MS/MS) was employed for metabolic analysis. The H2O2-induced oxidative damage was used in human embryonic kidney 293T (H293T) cells to assess intracellular antioxidant activities. Cells incubated with flower and fruit extracts showed better cell survival, lower levels of reactive oxygen species (ROS), and higher catalase and superoxide dismutase activities than those incubated with root, stem, and leaf extracts (p < 0.01). A total of 13 compounds were newly identified as intracellular antioxidants by association analysis, including coniferin, galactinol, trehalose, beta-D-lactose, trigonelline, nicotinamide-N-oxide, shikimic acid, 5′-deoxy-5′-(methylthio)adenosine, salicylic acid, isorhamnetin-3-O-neohespeidoside, methylhesperidin, 4-hydroxybenzoic acid, and cis-aconitic acid (R2 > 0.8, Log2FC > 1, distribution > 0.1%, and p < 0.01). They showed lower molecular weight and higher polarity, compared to previously identified in vitro antioxidants in D. nobile (p < 0.01). The credibility of HPLC-MS/MS relative quantification was verified by common methods. In conclusion, some saccharides and phenols with low molecular weight and high polarity helped protect H293T cells from oxidative damage by increasing the activities of intracellular antioxidant enzymes and reducing intracellular ROS levels. The results enriched the database of safe and effective intracellular antioxidants in medicinal plants.

Metabolic and Transcriptomic Profile Revealing the Differential Accumulating Mechanism in Different Parts of Dendrobium nobile

“…S1. The total ion chromatograms of flowers and fruits and the detailed identification information on metabolites are shown in Rao et al [6,7]. Detailed information on related metabolites is shown in Table S1.…”

“…Detailed information on related metabolites is shown in Table S1. [6,7]. Detailed information on related metabolites is shown in Table S1.…”

“…The solvent gradient was set as follows: 2% B, 1 min; 2-100% B, 18.0 min; 100% B, 22.0 min; 100-5% B, 22.1 min; and 5% B, 25 min. The QTRAP ® 6500+ mass spectrometer was operated in positive polarity mode with a curtain gas of 35 psi, a collision gas of medium, an ionspray voltage of −4500 V, a temperature of 500 • C, an ion source gas of 1:55, and an ion source gas of 2:55 [7].…”

“…To identify metabolites in the extracts of D. nobile quickly, accurately, and extensively, MRM was used for scanning mainly based on the above six key parameters [7,30]. For the Q1/Q3 scan, ± 0.7 was set; 0-300 was set for the DP scan; ± 150 was set for the CE scan; and ± 0.01 was set for the RT scan.…”

“…Alkaloids from D. nobile showed alleviating effects on neurotoxicity, synaptic deficits, and Alzheimer's disease-like symptoms [5]. Flavonoids and phenolic acids from D. nobile showed extracellular and intracellular antioxidant activities [6,7]. However, the systematic composition and distribution of all bioactive components in D. nobile were poorly reported.…”