Identification of the active compounds in the Yi-Fei-San-Jie formula using a comprehensive strategy based on cell extraction/UPLC-MS/MS, network pharmacology, and molecular biology techniques.

BACKGROUND: Chinese herbal formulae has multiple active constituents and targets, and the good clinical response is encouraging more scientists to explore the bio-active ingredients in such complex systems. Yi-Fei-San-Jie formula (YFSJF) is commonly used to treat patients with lung cancer in South China; however, its bio-active ingredients remain unknown. PURPOSE: We investigated the bio-active ingredients of the YFSJF using a novel comprehensive strategy. METHODS: A549 cell extraction coupled with ultra-high performance liquid chromatography-mass spectrometry (UPLC-MS/MS) was used for the screening of potential bio-active ingredients. Network pharmacology approach and molecular dynamics simulation were performed for the screening of targets. Surface plasmon resonance (SPR) assay and molecular biology techniques were used to verify the targets. RESULTS: Nine A549 cell membrane-binding compounds were identified through cell extraction/UPLC-MS/MS. Five compounds, namely ginsenoside Ro, ginsenoside Rb1, ginsenoside Rc, peimisine, and peimine were cytotoxic to A549 cells, and they were considered the bio-active ingredients of the YFSJF in vitro. Network pharmacology analysis revealed that TGFBR2 is the key target and the TGFß pathway is the key pathway targeted by YFSJF in non-small cell lung cancer. Peimisine showed an affinity to TGFBR2 using molecular docking and dynamic stimulation, which was confirmed using surface plasmon resonance spectroscopy. The molecular biology-based analysis further confirmed that peimisine targets TGFBR2 and can reverse A549 epithelial-mesenchymal transition by inhibiting the TGFß pathway. CONCLUSION: Taken together, cell extraction/UPLC-MS/MS, network pharmacology, and molecular biology-based analysis comprise a feasible strategy to explore active ingredients in YFSJF.

Identification of Active Ingredients of Huangqi Guizhi Wuwu Decoction for Promoting Nerve Function Recovery After Ischemic Stroke Using HT22 Live-Cell-Based Affinity Chromatography Combined with HPLC-MS/MS.

OBJECTIVE: The Chinese medicine Huangqi Guizhi Wuwu Decoction (HGWD) has been reported to improve the clinical symptoms and restore nerve function after ischemic stroke; however, its active ingredients are not well-determined. Therefore, this study aimed to investigate the bioactive compounds of HGWD and explore the possible mechanism of action. METHODS: The methods, including live HT22 cells, solid-phase extraction, and HPLC-MS/MS were utilized. The potential ingredients were identified through comparisons with literature and monomer compounds. Then, oxygen-glucose deprivation reperfusion (OGD/R)-treated HT22 cells were utilized to investigate the effect of HGWD components with specific binding affinities. Reactive oxygen species (ROS), superoxide dismutase (SOD), lactate dehydrogenase (LDH), and Tunel staining were used as testing indexes to analyze the protective effects of potential active ingredients on OGD/R-induced damage. RESULTS: Eleven compounds with specific binding affinities were identified as calycosin-7-O-glucoside, calycosin, formononetin, cinnamic alcohol, cinnamic acid, betaine, dl-2-phenylpropionic acid, 4-hydroxycinnamic acid, 6-methylcoumarin, wogonin, and paeoniflorin. Among them, six compounds had a protective effect on OGD/R-treated HT22 cells. Furthermore, calycosin-7-O-glucoside, calycosin, paeoniflorin, 4-hydroxycinnamic acid, wogonin, and formononetin could regulate oxidative stress and apoptosis to attenuate the cell damage caused by OGD/R. CONCLUSION: The mechanism of action of HGWD to promote neurological recovery after ischemic stroke was related to the regulation of oxidative stress and apoptosis. This study suggested that cell membrane affinity chromatography combined with HPLC-MS/MS could be applied to screen potential active components in traditional Chinese medicines (TCM).

A silver nanorod resonance rayleigh scattering-energy transfer analytical platform for trace tea polyphenols.

The stable silver nanorod (AgNR) sol in red was prepared by the two-step procedure of NaBH4-H2O2 and citrate heating reduction, and it exhibited a strong resonance Rayleigh scattering (RRS) peak at 346 nm. In pH 3.8 HAc-NaAc buffer solution, tea polyphenols (TP) reacted with ammonium molybdate (AM) to form yellow organic molybdate (OM) as receptor that was closed to the donor of AgNR, the RRS energy transfer (RRS-ET) takes place, owing to the overlapping between the AgNR RRS spectra and OM absorption spectra. When TP concentration increased, the RRS intensity decreased due to the RRS-ET increasing. So, a simple and sensitive AgNR surface plasmon RRS-ET analytical platform was fabricated to detect trace TP in the range of 0.05-0.85 µg/mL, with a detection limit of 0.03 µg/mL TP. The TP in tea samples was analyzed by this RRS-ET analysis platform, with satisfactory results.

[Fluorescence Determination of Trace Se with the Hydride-K13-Rhodamine 6G System].

Se is a necessary trace element for human and animals, but the excess intake of Se caused poison. Thus, it is very important to determination of Se in foods and water. The target of this study is development of a new, sensitive and selective hydride generation-molecular fluorescence method for the determination of Se. In 0. 36 mol . L-1 sulfuric acid, NaBH4 as reducing agent, Se (IV) is reduced to H2 Se. Usin3-g I solution as absorption liquid3, I- is reduced to I- by H2Se. When adding rhodamine 6G, Rhodamine 6G and I3- form association particles, which lead to the fluorescence intensity decreased. When Se(IV) existing, Rhodamine 6G and I3- bind less, And the remaining amount of Rhodamine 6G increase. So the fluorescence intensity is enhanced. The analytical conditions were optimized, a 0. 36 ml . L-1 H2SO4, 21. 6.g . L-1 NaBH4, 23.3 µm . L-1 rhodamine 6G, and 50 µmol . L-1 KI3 were chosen for use. When the excitation wavelength is at 480nm, the Rayleigh scattering peak does not affect the fluorescence recording, and was selected for determination of Se. Under the selected conditions, Se(IV) concentration in the 0. 02~0. 60 µg . mL-1 range and the increase value of the fluorescence intensity (ΔF) at 562 nm linear relationship. The linear regression equation is ΔF562 nm =12. 6c + 20. 9. The detecton limit was 0.01 µ.g . L-1. The influence of coexistence substances on the hydride generatin-molecular fluorescence determination of 5. 07 X10(-6) mol . L-1 Se(IV) was considered in details. Results showed that this new fluorescence method is of high selectivity, that is, 0. 5 mmol. L-1 Ba2+, Ca2+, Zn2+ and Fe3+, 0. 25 mmol . L-1 . Mg2+, 0. 05 mmol . L-1 K+, 0. 2 mmol . L-1 Al3+, 0. 025 mmol . L-1 Te(VI) do not interfere with the determination. The influence of Hg2+, CD2+ and Cu2+ that precipitate with Se(IV), can be eliminated by addition of complex reagent. This hydride generation-molecular fluorescence method has been applied to determination of trace Se in water samples,

[Studies on flavones in the leaves of Cyclocarya paliums (batal.) Iljinsk].

To do investigation of flavones from the leaves of Cyclocarya paliums (Batal.) Iljinsk. The pure products were isolated by column chromatography of silica gel, thin layer chromatography of silica gel and column chromatography of Sephadex LH-20, their chemical structures were elucidated by NMR, IR and UV spectroscopic evidence. Four flavones were isolated from the leaves of Cyclocarya paliums. They are quercetin (I), kaempferol (II), kaempferol-7-O-alpha-L-rhamnopyranosyl (III) ,and kaempferol-4'-methoxyl-7-O-beta-D-mancopyranosyl (IV). Compounds III and IV are obtained for the first time from this plants.