Anti-inflammatory effects of anthraquinones of Polygonum multiflorum roots

Abstract

Background: The roots of Polygonum multiflorum Thunberg (Polygonaceae) are a crude drug known as Kashu and are thought to improve blood deficiency as a Kampo concept. Kashu is added to Kampo formulas, including Tokiinshi, which is used to treat eczema and dermatitis with itchiness by facilitating blood circulation in the skin. Although anti- inflammatory effects were expected, the reports on anti-inflammatory constituents of Polygonum roots are very few. Objective: To purify pharmacologically active constituents from Polygonum roots and compare their anti- inflammatory potency using hepatocytes. Methods: Polygonum roots were extracted and fractionated based on hydrophobicity to three fractions, the ethyl acetate (EtOAc)-soluble, n-butanol-soluble, and water-soluble fractions. Constituents were purified from the EtOAc- soluble fraction, and its contents were measured using liquid chromatography-triple quadrupole mass spectrometer. The interleukin (IL)-1β-induced production of nitric oxide (NO), a proinflammatory mediator, in rat hepatocytes was monitored to assess anti-inflammatory activity. Results: The EtOAc-soluble fraction inhibited IL-1β-induced NO production, whereas the other fractions did not. Six constituents were isolated from the EtOAc-soluble fraction and identified as (E)-2,3,5,4'-tetrahydroxystilbene-2-O-β- D-glucoside, emodin, emodin-8-O-β-D-glucopyranoside, physcion, physcion-8-O-β-D-glucopyranoside, and catechin. Emodin and emodin-8-O-β-D-glucopyranoside significantly inhibited NO production in hepatocytes, and the others showed less potency in suppressing NO production. These two anthraquinones decreased the protein and mRNA levels of inducible NO synthase, as well as the mRNA levels of tumor necrosis factor α. In a search of the literature to date, the comparison of pharmacological activity of the constituents of Polygonum roots was not reported.Conclusion: These results suggest that hydrophobic constituents in the EtOAc-soluble fraction, especially anthraquinones, may exert the anti-inflammatory effects by treatment with Polygonum roots.Keywords: anthraquinone, nitric oxide, inflammation, Kampo medicine, LC–MS/MS.

Identification of Anti-Inflammatory Compounds from Peucedanum praeruptorum Roots by Using Nitric Oxide-Producing Rat Hepatocytes Stimulated by Interleukin 1β

“…After 4 h incubation of hepatocytes with 1 nM rat IL-1β ± a compound on day 1, total RNA was prepared from hepatocytes using Sepasol I Super G solution (Nacalai Tesque, Inc.) [ 4 , 8 ]. The cDNA was reverse-transcribed from total RNA and amplified by PCR with the primers described in Table 3 .…”

“…According to the ΔΔCt method, the relative mRNA levels were calculated from these obtained Ct values. The Ct values were normalized to EF-1α mRNA [ 4 , 8 ], which is encoded by a housekeeping gene (an internal control). The normalized mRNA levels in the total RNA from IL-1β-treated hepatocytes were set at 100%.…”

“…When NO production in the presence of a drug is compared with that in the presence of IL-1β alone, the potency of NO suppression can be calculated and expressed as a half-maximal inhibitory concentration (IC 50 ) value [ 3 , 5 ]. Principal constituents in a crude drug have been identified by comparing the IC 50 values of the compounds [ 6 , 7 , 8 ]. The proinflammatory genes include iNOS, proinflammatory cytokine, and chemokine genes, most of which are regulated by transcription factor NF-κB [ 9 ].…”