Identifying Anti-NSCLC Bioactive Compounds in Scutellaria via 2D NMR-Based Metabolomic Analysis of Pharmacologically Classified Crude Extracts.

We presented a strategy utilizing 2D NMR-based metabolomic analysis of crude extracts, categorized by different pharmacological activities, to rapidly identify the primary bioactive components of TCM. It was applied to identify the potential bioactive components from Scutellaria crude extracts that exhibit anti-non-small cell lung cancer (anti-NSCLC) activity. Four Scutellaria species were chosen as the study subjects because of their close phylogenetic relationship, but their crude extracts exhibit significantly different anti-NSCLC activity. Cell proliferation assay was used to assess the anti-NSCLC activity of four species of Scutellaria. 1H-13C HSQC spectra were acquired for the chemical profiling of these crude extracts. Based on the pharmacological classification (PCA, OPLS-DA and univariate hypothesis test) were performed to identify the bioactive constituents in Scutellaria associated with the anti-NSCLC activity. As a result, three compounds, baicalein, wogonin and scutellarin were identified as bioactive compounds. The anti-NSCLC activity of the three potential active compounds were further confirmed via cell proliferation assay. The mechanism of the anti-NSCLC activity by these active constituents was further explored via flow cytometry and western blot analyses. This study demonstrated 2D NMR-based metabolomic analysis of pharmacologically classified crude extracts to be an efficient approach to the identification of active components of herbal medicine.

Effect of Different Processing Methods on the Chemical Constituents of Scrophulariae Radix as Revealed by 2D NMR-Based Metabolomics.

Scrophulariae Radix (SR) is one of the oldest and most frequently used Chinese herbs for oriental medicine in China. Before clinical use, the SR should be processed using different methods after harvest, such as steaming, "sweating", and traditional fire-drying. In order to investigate the difference in chemical constituents using different processing methods, the two-dimensional (2D) 1H-13C heteronuclear single quantum correlation (1H-13C HSQC)-based metabolomics approach was applied to extensively characterize the difference in the chemical components in the extracts of SR processed using different processing methods. In total, 20 compounds were identified as potential chemical markers that changed significantly with different steaming durations. Seven compounds can be used as potential chemical markers to differentiate processing by sweating, hot-air drying, and steaming for 4 h. These findings could elucidate the change of chemical constituents of the processed SR and provide a guide for the processing. In addition, our protocol may represent a general approach to characterizing chemical compounds of traditional Chinese medicine (TCM) and therefore might be considered as a promising approach to exploring the scientific basis of traditional processing of TCM.

[Study on HPLC fingerprint of Sabia parviflora].

This research was performed to establish the HPLC fingerprint of Sabia parviflora. HPLC method was carried out on a Thermo Accucore-C18(4. 6 mm×150 mm,2. 6 µm) column by 30% tetrahydrofuran in methyl alcohol-acetonitrile-0. 1% phosphate solution as mobile phase at a flow rate of 1. 0 m L·min-1,the column temperature was 30 ℃ and the detection wavelength was 360 nm. The fingerprints were further evaluated by chemometrics methods including similarity analysis,hierarchical clustering analysis,and principal component analysis. In HPLC fingerprint,15 common peaks were selected as the common peaks,and 6 contents of them were identified. The similarity degrees of 38 batches of the samples was more than 0. 710,and the samples were divided into 6 clusters by their quality difference. The method was precision,repeatable,stable,simple and reliable,which could be used for quality control and evaluation of S. parviflora.

Anti-proliferative effect of RCE-4 from Reineckia carnea on human cervical cancer HeLa cells by inhibiting the PI3K/Akt/mTOR signaling pathway and NF-κB activation.

Cervical cancer is the second leading cause of cancer deaths in women worldwide. In recent years, the studies find that inflammation is a critical component of tumor progression, and the ideal therapeutic methods should be aimed at the inflammation reaction triggers. (1ß,3ß,5ß,25S)-spirostan-1,3-diol1-[α-L-rhamnopyranosyl-(1 → 2)-ß-D-xylopyranoside] (RCE-4) was the main active composition of Reineckia carnea (Andr.) Kunth. It significantly induced apoptosis in cervical cancer Caski cells through the mitochondrial pathway in our previous studies; however, its underlying mechanism remains poorly understood. This study aimed to further evaluate the effect of RCE-4 on human cervical cancer HeLa cells. Based on this observation, we investigated the anti-cervical cancer effect of RCE-4 by modulating phosphatidylinositol 3-kinase/protein kinase-B/mammalian target of rapamycin (PI3K/Akt/mTOR) signaling pathway, nuclear factor-kappa B (NF-κB) activation, and inflammation-related key factors in HeLa cells. The results indicated that the HeLa cell was the most sensitive with an IC50 of 7.01 µM; RCE-4 significantly promoted the release of cellular lactate dehydrogenase (LDH); increased DNA fragmentation and apoptosis; reduced PI3K, Akt, mTOR, and NF-κBp65 phosphorylation levels; increased the Bax and cleaved poly (ADP-ribose) polymerase (PARP) protein levels; suppressed Bcl-2 protein expression; elevated the Bax/Bcl-2 expression ratio; and decreased the interleukin-1 beta (IL-1ß) and interleukin-6 (IL-6) mRNA expressions in HeLa cells in a concentration-dependent manner. These findings suggest that RCE-4 exerted beneficially anti-cervical cancer effect on HeLa cells, mainly inhibiting PI3K/Akt/mTOR signaling pathway phosphorylation and NF-κB activation, promoting HeLa cell apoptosis. Graphical abstract Anti-tumor effect of RCE-4 on HeLa cells.

Protective effect of saponins extract from Panax japonicus on myocardial infarction: involvement of NF-κB, Sirt1 and mitogen-activated protein kinase signalling pathways and inhibition of inflammation.

OBJECTIVE: Inflammation is widely acknowledged to increase morbidity and mortality in myocardial infarction (MI), and the ideal therapeutic methods should be aimed at the inflammation reaction triggers. The aim was to evaluate the protective effect of saponins extracted from Panax japonicus (SPJ) on MI, and based on these results investigate the possible involvement mechanism of the nuclear factor-kappa B (NF-κB), sirtuin1 (SIRT1) and mitogen-activated protein kinases (MAPKs) signalling pathways. METHODS: Sprague-Dawley rats were randomly divided into sham, MI, MI + SPJ 50 and SPJ 100 mg/kg groups. After administration for 3 days, MI rats were created by ligaturing coronary artery, and then underwent the same administration for 7 days as before. Cardiac function and the expressions of pro-apoptosis protein Bax, anti-apoptosis protein Bcl-2, NF-κB, SIRT1, MAPKs signal pathway-related proteins and inflammatory factor, such as tumour necrosis factor alpha (TNF-α) and monocyte chemoattractant protein 1 (MCP-1), were assessed. KEY FINDINGS: SPJ might significantly improve cardiac function, decrease the serum MCP-1 and TNF-α levels, ameliorate the increased Bax protein expression and decrease Bcl-2 protein expression, and suppress the protein expressions of NF-κBp65 subunit, extracellular signal-regulated kinase 1/2 (ERK1/2) and p38 MAPK, but have no effect on c-Jun NH2-terminal kinase, and increase the expression of SIRT1. Histopathological observations provided supportive evidence for aforementioned results, and with the dose of SPJ increasing, the aforesaid improvement became more and more strong. CONCLUSION: The studies demonstrated that SPJ exerted beneficially cardioprotective effects on MI rats, mainly inhibiting NF-κB, ERK1/2 and p38 MAPK activation, but increased the expression of SIRT1, alleviating MI injury and cardiac cell death.