Comprehensive evaluation and anti-inflammatory activity of “Zhi” polygonatum cyrtonema produced by the classical steaming approach

Abstract

IntroductionPolygonati Rhizoma, known as “Huangjing”, was first documented in the well-known book Mingyi Bielu, written in the Chinese Han Dynasty. A "Nine-Steam-Nine-Bask" method is the best way to increase their tonic function by steaming and drying the raw rhizomes. This study was conducted with the primary objective of assessing "Zhi" P. cyrtonema qualitatively and quantitatively. Additionally, in vitro studies of the anti-inflammatory activity of "Zhi" P. cyrtonema revealed its pharmacodynamic effects and possible mechanisms of action. MethodsP. cyrtonema was steamed and dried nine times and subsequently subjected to hot water extraction and ethanol precipitation to obtain the desired P. cyrtonema polysaccharides (PCPs). Several approaches, including HPGPC-RID, NMR, and GC-MS, were used to evaluate the molecular weight, composition changes, and monosaccharide composition of PCPs during steaming. Multivariate data analysis, including PCA and HCA, was applied for the divergence analysis of "Zhi" P. cyrtonema. The anti-inflammatory effect of 4-Z PCPs on MH7A cells was evaluated using a cell viability assay. ELISA assays, DAPI staining, cell cycle assay, and scratching experiments revealed the potential mechanism. ResultsWith increasing steaming time, the color of the herb gradually changes from yellow to black and finally to dark black. In processed roots, the levels of polysaccharides decreased, while monosaccharides, such as galactose and glucose, increased. The high-molecular-weight polysaccharide was partially hydrolyzed, forming smaller polysaccharides, monosaccharides, and trace oligosaccharides. In the HPGPC profile, peak 1, with a retention time of 5.20 min and a molecular mass of 1140.51 kDa, can transform into peak 2 (6.10 min, 374.00 kDa) and peak 3 (9.71 min, 9.73 kDa). Using NMR analysis, it has been shown that 5-hydroxymethylfurfural (5-HMF), a toxic substance formed by the thermal degradation of saccharides, is present from the start of the fifth processed herb but is rare before 5-Z P. cyrtonema. The GC-MS analysis also revealed the presence of various monosaccharides, including mannose, galactose, glucose, fructose, rhamnose, arabinose, and galacturonic acid in "Zhi" P. cyrtonema. Using saccharides as variables, we were able to classify "Zhi" P. cyrtonema into four groups: fresh and 1-Z PCPs in groups A, 2-Z, 3-Z, 4-Z, 5-Z, 6-Z, 7-Z and 8-Z PCPs in group C, and 9-Z PCPs in group D. Results of the MH7A cell experiment indicate that 4-Z PCPs are more efficient than 0-Z and 9-Z PCP. After the fourth processing cycle, the differences between metabolic profiles became smaller. ConclusionsA combination of UV–Vis spectra, HPGPC-RID analysis, and NMR techniques were used in this study to achieve qualitative and quantitative analyses of "Zhi" PCPs, and we were able to make a comprehensive evaluation of "Zhi" P. cyrtonema. Meanwhile, using saccharides as variables by the HCA and PCA method, we were able to divide "Zhi" P. cyrtonema into four groups, namely fresh and 1-Z PCPs as group A, and 2-Z, 3-Z and 4-Z PCPs as group B, as well as 5-Z, 6-Z, 7-Z, 8-Z as group C, and 9-Z PCPs as group D. The anti-inflammatory activity studies indicated that the 4-Z PCPs better inhibited the growth of MH7A cells and induced apoptosis of synovial fibroblasts, suggesting that the 4-Z PCPs could be used as an anti-inflammatory reagent.