Preparative isolation of antioxidative furanocoumarins from Dracocephalum heterophyllum and their potential action target.

Traditional Tibetan medicine has extensively documented the health benefits of Dracocephalum heterophyllum. However, there are few reports on the chemical composition of furanocoumarins, probably because of their complicated isolation and purification procedures. In this study, four antioxidative furanocoumarins were isolated from Dracocephalum heterophyllum by medium- and high-pressure liquid chromatography in combination with on-line high-performance liquid chromatography-1,1-diphenyl-2-picrylhydrazyl recognition. Crude samples were sequentially pretreated by medium-pressure liquid chromatography using silica gel, MCI GEL CHP20P, and diol as stationary phases, whereas on-line high-performance liquid chromatography-1,1-diphenyl-2-picrylhydrazyl system was used to recognize antioxidant peaks in target fractions. Thereafter, the antioxidative peaks were separated and purified through high-pressure liquid chromatography to obtain four furanocoumarins with purities greater than 95%; namely isodemethylfuropinarine, demethylfuropinarine, alloimperatorin, and alloisoimperatorin. Finally, the antioxidant capacity of the isolated furanocoumarins was determined using in vitro experiments (1,1-diphenyl-2-picrylhydrazyl assays, molecular docking, and cellular validation) and it was concluded that nuclear factor erythroid 2-related factor 2 protein is a potential target of these compounds for their antioxidation effects. Thus, the proposed methodology exhibits excellent efficacy for the preparative isolation of high-purity antioxidative furanocoumarins from extracts of Dracocephalum heterophyllum and it can be efficiently utilized for isolating antioxidants from other natural products.

The Isolation and Preparation of Samwinol from Dracocephalum heterophyllum and Prevention on Aß25-35-Induced Neuroinflammation in PC-12 Cells.

Dracocephalum heterophyllum (D. heterophyllum) is a traditional Chinese Tibetan medicine that has been used for the treatment of lymphitis, hepatitis, and bronchitis. However, only a few selected chemical components are currently obtained from D. heterophyllum, which limits its further pharmacological applications. In this study, we have obtained samwinol from D. heterophyllum by medium- and high-pressure liquid chromatography separation for the first time. Thereafter, we investigated the protective actions of samwinol against amyloid beta protein fragment 25-35 (Aß25-35) induced neurotoxicity in cultured rat pheochromocytoma PC-12 cells and explored its underlying mechanisms of action. The results indicated that samwinol could increase cell viability and inhibit the production of reactive oxygen species (ROS) and mitochondria-derived ROS, as assessed by MTT assay, Giemsa staining, and flow cytometry assay. Through Western blot analysis, it was found that samwinol substantially inhibited the phosphorylation of ERK(1/2) and promoted the expression of HO-1 and Nrf2. The data obtained from molecular docking were also consistent with the above conclusions. All of these results showed that samwinol from D. heterophyllum can display significant anti-neuroinflammatory and antioxidant activities in vitro, which are associated with the suppression of ERK/AKT phosphorylation and the activation of the Nrf2/HO-1 signaling pathway. In the future, additional in-depth mechanism studies will be carried out to provide more evidence for the potential of samwinol in the treatment of Alzheimer's disease.

Integrated chromatographic approach for the discovery of gingerol antioxidants from Dracocephalum heterophyllum and their potential targets.

As a traditional Tibetan medicine, Dracocephalum heterophyllum has many benefits, but due to the complicated procedures of separation and purification of its chemical constituents, there are few reports on gingerols. In this study, four antioxidative gingerols were isolated from Dracocephalum heterophyllum by an integrated chromatographic approach. Antioxidant activity was then determined by in vitro experiments and its potential targets of action were investigated. First, the extract was pretreated using silica gel, MCI GEL®CHP20P, and diol and spherical medium pressure columns, while the antioxidant peaks were identified using an online HPLC-1,1-diphenyl-2-picrylhydrazyl system. Then, the antioxidant peaks were directionally separated and purified by high pressure liquid chromatography to obtain four gingerols with a purity higher than 95%, namely 5-methoxy-6-gingerol, 6-shogaol, 6-paradol, and diacetoxy-6-gingerdiol. Finally, 1,1-diphenyl-2-picrylhydrazyl assays and cellular antioxidant experiments were carried out, and molecular docking was used to explore potential antioxidant targets. The isolated gingerols upregulated the activity of antioxidant enzymes, including superoxide dismutase (SOD), heme oxygenase-1 (HO-1) and NADPH oxidase 2 (NOX2), while they had little effect on the activity of nadph:quinone oxidoreductase-1 (NQO1). This method can efficiently prepare and isolate antioxidative gingerols from Dracocephalum heterophyllum, and it can be extended to isolate antioxidants from other natural products.