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2 protocols using baicalein

1

Profiling Phytochemical Composition of Skullcap

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The dried and pulverised biomass samples were extracted with 50 mL of analytical-grade methanol for 2 h at 78 ± 2 °C under a reflux condenser. After extraction, the obtained extracts were filtered through a paper filter, transferred to crystallisers, and left at room temperature to allow the solvent to evaporate. The remains were dissolved in 4 mL of HPLC-grade methanol (Merck) and subjected to HPLC. Qualitative and quantitative HPLC with diode array detection (DAD) was performed as described previously [20 (link),66 (link)]. Compounds were identified based on their retention times and UV spectra, as well as by comparison with reference substances (27 flavonoids, 19 phenolic acids, benzoic and cinnamic acids, 2 phenylethanoid glycosides). The reference standards used for the HPLC analyses of skullcap-specific flavonoids were baicalein, baicalin, scutellarin, wogonin (ChromaDex, Irvine, CA, USA), chrysin, scutellarein, wogonoside (Sigma–Aldrich Co., Saint Louis, MO, USA), oroxylin A and skullcapflavone II (ChemFaces Biochemical Co., Wuhan, China). The verbascoside was purchased form ChromaDex, Irvine, CA, USA.
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2

Neurochemical Signaling Pathway Analysis

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Methanol, hexane, chloroform, ethyl acetate, butanol, lipopolysaccharide (LPS) from Escherichia coli serotype 026:B6 (Lot# 021M4072V), (-)-nicotine, methyllycaconitine citrate salt hydrate (MLA) from Delphinium brownii seeds, mecamylamine hydrochloride and 7-hydroxy-3H-phenoxazin-3-one-10-oxide sodium salt (resazurin) were purchased from Sigma-Aldrich (St Louis, MO, USA). Astragalin, baicalein, catechin, daidzin, daidzein, delphinidin, genistein, isoquercitrin, isorhamnetin, malvidin, petunidin-3-glycoside, quercetagetin, quercetin, rhamnetin, sakuranetin, spiraeoside and tamarixetin (Fig. 1) were purchased from Chromadex (Irvine, CA, USA). [3H]-MLA (∼60Ci/mmol), [3H]-cytisine (∼16Ci/mmol) and [3H]-epibatidine (∼30Ci/mmol) were purchased from American Radiolabeled Chemicals, Inc. (St. Louis, MO, USA). Antibiotics (10000U/mL penicillin and 10000ug/mL streptomycin), 2.5% trypsin (10×), fetal bovine serum (FBS), Dulbeccos' Modified Eagle Medium (DMEM), DMEM:Nutrient mixture F-12 (DMEM/F-12) and Hanks Balanced Salt Solution (HBSS) were purchased from Life Technologies Corporation (Grand Island, NY, USA). Griess reagent system was purchased from Promega Corporation (Madison, WI, USA). TNF-alpha ELISA Ready-SET-Go!® was purchased from eBioscience Inc. (San Diego, CA, USA).
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