Abstract
Ethnopharmacological relevanceFlowers from Styrax japonicus sieb. et Zucc. have been used as a Chinese folk medicine to alleviate pain such as toothache and sore throat. Aim of the studyTo testify the analgesic effect of flowers from Styrax japonicus, analyze components of the active fraction, and investigate the mechanism of analgesia. Materials and methodsFlower extracts were obtained by ethanol, petroleum ether and hydrodistillation extraction. Different fractions of ethanol extracts (EE) were isolated by silica gel column chromatography and preparative liquid chromatography. Analgesic effects of EE, petroleum ether extracts (PEE), hydrodistillation extracts (HDE), and fractions of EE were evaluated using hot plate, acetic acid-induced writhing and formalin tests on mice. Components of the active fraction 1 (F1) were determined by the ultrahigh-performance liquid chromatography Q extractive mass spectrometry (UHPLC-QE-MS). Anti-inflammatory and sedative effects involving analgesic mechanisms were evaluated by carrageenan induced hind paw oedema and pentobarbital sodium sleep tests, respectively. In addition, antagonists including naloxone hydrochloride (NXH), flumazenil (FM), SCH23390 (SCH) and WAY100635 (WAY) were used to investigate the possible mechanism of analgesia. Contents of neurotransmitters and relevant metabolites in different brain regions of mice were also quantified by the ultraperformance liquid chromatography with a fluorescence detector (UPLC-FLD). ResultsEE rather than PEE and HDE at medium and high doses (150 mg/kg and 300 mg/kg) significantly prolonged the latency time of the response of mice to the thermal stimulation in the hot plate test. Moreover, EE significantly decreased number of writhes in the acetic acid-induced writhing test, and reduced licking time in both two phases of the formalin test in a dose-dependent manner. The F1 (50 mg/kg) showed effective antinociceptive responses in all mice models. However, fraction 2 (F2) and fraction 3 (F3) at 50 mg/kg performed no analgesic action. Kaempferol-3-O-rutinoside, isorhamnetin-3-O-rutinoside, pinoresinol-4-O-glucoside, forsythin and arctiin were identified from components of the F1. Furthermore, F1 (50 mg/kg) did not significantly affect hind paw oedema of mice induced by carrageenan but significantly shortened sleep latency and increased sleep duration in the pentobarbital sodium sleep test. In addition, the antinociceptive response of F1 was not affected by NXH in two mice models, but significantly blocked by FM and WAY in the hot plate test. In the formalin test, FM avoided the effect of F1 only in the first phase, while the analgesic activity of F1 was totally suppressed by WAY in both two phases. Otherwise, contents of 5-hydroxytryptamine (5-HT) and 5-hydroxyindoleacetic acid (5-HIAA) increased significantly in hippocampus and striatum of mice in the F1 group. ConclusionEE from flowers of Styrax japonicus, and F1, the active part isolated from EE, showed significant antinociceptive activities. The analgesic effect of F1 appeared to be related to the sedative effect, partially mediated by the GABAergic system, and highly involved in the serotonergic system. This was the first study confirming the analgesic effect of Styrax japonicus flower, which provided a candidate for the development of non-opioid analgesics.
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