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Aerial parts, leaves, and stems of  Gaultheria procumbens  are polyphenol-rich herbal medicines with anti-inflammatory and antioxidant effects. The present study focused on identifying active markers of the  G. procumbens  extracts in an integrated approach combining phytochemical and biological capacity tests. The target compounds, representing all classes of  Gaultheria  polyphenols, were pre-selected by LC-ESI-PDA-MS/MS. For unambiguous identification, the key analytes, including a rare procyanidin trimer (cinnamtannin B-1), miquelianin potassium salt, and two new natural products: quercetin and kaempferol 3- O - β - d -xylopyranosyl-(1→2)- β - d -glucuronopyranosides, were isolated by preparative HPLC and investigated by spectroscopy (HR-ESI-MS, UV-vis, CD, 1D- and 2D-NMR), thiolysis, flame photometry, optical rotation experiments, and absolute configuration studies. The significant contribution of the pre-selected compounds to the biological effects of the extracts was confirmed in vitro: the analytes significantly and in a dose-dependent manner down-regulated the pro-oxidant and pro-inflammatory functions of human neutrophils ex vivo (inhibited the release of reactive oxygen species, IL-1β, TNF-α, and neutrophils elastase, ELA-2), inhibited two key pro-inflammatory enzymes (cyclooxygenase, COX-2, and hyaluronidase), and most of them, except gaultherin, exerted potent direct antioxidant activity (ferric reducing antioxidant power and superoxide anion scavenging capacity). Moreover, cellular safety was confirmed for all compounds by flow cytometry. Eventually, as these mechanisms have been connected to the health benefits of  G. procumbens , 11 polyphenols were accepted as active markers, and a simple, accurate, reproducible, and fully validated RP-HPLC-PDA method for standardisation of the target extracts was proposed.

international journal of molecular sciences

Screening for the Active Anti-Inflammatory and Antioxidant Polyphenols of Gaultheria procumbens and Their Application for Standardisation: From Identification through Cellular Studies to Quantitative Determination