Screening nephroprotective compounds from cortex Moutan by mesangial cell extraction and UPLC

A method for screening nephroprotective compounds in cortex Moutan, a common traditional Chinese medicine (TCM) in treating diabetic nephropathy with renal mesangial cell extraction and ultra performance liquid chromatography technique was described in this paper. We hypothesize that the compounds which bind to cell membranes under pathological conditions may be the bioactive compounds in TCMs. Mesangial cells were cultured in medium containing 5 mM (physiological, NG) or 30 mM (pathological, HG) glucose for 48 h and then incubated with cortex Moutan extract. After the unbound substances were washed off, the cell membrane-bound compounds were dissociated and concentrated by an SPE column. By comparing the chromatograms of NG and HG cultured-cell extractions and cortex Moutan extract, three compounds bound to both NG and HG-cultured mesangial cells were identified as paeoniflorin, pentagalloylglucose (PGG) and paeonol. In vitro studies showed that paeoniflorin, PGG and paeonol reduced the activity of nicotinamide-adenine dinucleotide phosphate oxidase (NADPH) activity, and decreased the level of reactive oxygen species (ROS), transforming growth factor-β1 (TGF-β1) and fibronectin in high glucose cultured mesangial cells. The results indicate that paeonol, paeoniflorin and PGG may be the nephroprotective compounds from cortex Moutan. This study is expected to provide a more reliable and effective method for screening bioactive compounds from the complex TCM systems.