Gua-Lou-Gui-Zhi decoction (GLGZD) is a classical multiherb traditional Chinese medicine formula that has ameliorative effects on oxygen-glucose deprivation/reperfusion (OGD/R) injury and has been applied for the treatment of stroke in clinical practice; however, its active ingredients remain unknown. The aim of this study was to develop an effective method for screening for components of GLGZD with potential therapeutic activity against OGD/R injury. Brain microvascular endothelial cell membrane-coated magnetic beads (CMs@rBMECs-MBs) were incubated with the GLGZD extract; the bound material was eluted and the constituents were identified using solid phase extraction and ultra-performance liquid chromatography-Orbitrap Fusion Tribrid mass spectrometry (UPLC-Orbitrap Fusion Tribrid MS). The biological activities of the identified GLGZD components were analyzed using OGD/R-exposed brain endothelial cells. Seven compounds bound to the CMs@rBMECs-MBs were identified as gallic acid, paeoniflorin, liquiritigenin, isoliquiritigenin, liquiritin, isoliquiritin, and formononetin. Among them, six (except formononetin) protected brain endothelial cells against OGD/R injury in a concentration-dependent manner (20-120 μM; P < 0.01-0.05) and downregulated the expression of hypoxia-inducible factor-1α (P < 0.01) involved in the pathogenic mechanisms triggered by stroke. Our findings suggest that the screening of bioactive compounds using cell membrane-coated magnetic beads combined with solid phase extraction and UPLC-Orbitrap Fusion Tribrid MS is an effective method for the bio-specific extraction and identification of ingredients responsible for the therapeutic activity of traditional Chinese medicines.
Keywords: Cell membrane-coated magnetic beads; Gua-Lou-Gui-Zhi-decoction; Hypoxia-inducible factor-1α; Mass spectrometry; Oxygen-glucose deprivation/reperfusion injury; Stroke.
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