Bioactivity‐based ultra‐performance liquid chromatography‐coupled quadrupole time‐of‐flight mass spectrometry for NF‐κB inhibitors identification in Chinese Medicinal Preparation Bufei Granule

Abstract

Traditional Chinese medicine (TCM) preparations have become effective treatments for many diseases. However, their active ingredients are still uncertain and difficult to identify. In this study, we propose a strategy that integrates ultra-performance liquid chromatography/quadrupole-time-of-flight mass spectrometry (UPLC/Q-TOF-MS) and bioactive (NF-κB inhibitor) luciferase reporter assay systems for the rapid determination of various anti-inflammatory compounds of TCM preparations. In this way, Bufei Granule (BFG), a TCM preparation used for the clinical therapy of asthma, was analyzed by the two ways of component identification and activity detection. Potential anti-inflammatory constituents were screened by NF-κB activity assay systems and simultaneously identified according to the mass spectrometry data. Three structural types of NF-κB inhibitors (caffeic acid derivatives, flavonoids and Pentacyclic triterpenes) were characterized. Further cytokine detection confirmed the anti-inflammatory effects of the potential NF-κB inhibitors. Compared with conventional chromatographic separation and inhibitory activity detection, integrating UPLC/Q-TOF-MS identification and virtual validation was more convenient and more reliable. This strategy clearly demonstrates that MS data-based fingerprinting is a meaningful tool not only in identifying constituents in complex matrix but also in directly screening for powerful trace ingredients in TCM preparations. Copyright © 2016 John Wiley & Sons, Ltd.

An Optimized MicroPET Imaging Method for the Distribution and Synergies of Natural Products

“…GA and [ 18 F]-GA (100 μmol/L) were added to HEK 293T which has been stimulated by TNF-α. The activities were evaluated using the luciferase reporter assay system ( Qingxin et al, 2016 ). Dex (10 μmol/L) and GA showed significant NF-κB inhibitory effects ( P < 0.05), as shown in Figure 1 .…”

“…[ 18 F]-GA was collected into a plate and evaporated to dryness in a vacuum drying oven at 40°C. After 48 h, radioactive elements had decayed, the residues were stored at 4°C for anti-inflammatory activity tests ( Dong et al, 2013 ; Qingxin et al, 2016 ).…”

“…HEK 293 T cells were pretreated with drugs 1 h prior to TNF-stimulation (5 ng/mL) for 6 h. Dex (10 -5 mol/L) was used as a positive control for NF-κB inhibition. The NF-κB activity was assayed using dual luciferase reporter assay system (Promega) according to the manufacturer’s instructions ( Qingxin et al, 2016 ). Antiinflammatory activity was calculated as the ratio of firefly luciferase activity to Renilla luciferase activity (internal control).…”

Combining UPLC/Q-TOF-MS/MS With Biological Evaluation for NF-κB Inhibitors in Uyghur Medicine Althaea rosea Flowers

“…This method is sensitive, and the detailed method validation is complete. Information on potential inhibitors provided here will facilitate the development of new drugs and the development of plant resources (Qingxin et al, 2016). We demonstrate here that this integrated system has advantages in separation, structural identification, and bioactivity analysis of hit compounds (Halai and Cooper, 2012; Dong et al, 2013).…”

Biodistribution of andrographolide to assess the interior‐exterior relationship between the lung and intestine using microPET

“…The anti‐inflammatory activity of [ 19 F]‐AG (F‐AG) was determined according to previously reported methods 24 and calculated as the ratio of firefly luciferase activity to Renilla luciferase activity (internal control).…”