scopoletin and Inflammation

This protocol describes microsphere-based protease assays for use in flow cytometry and high-throughput screening. This platform measures a loss of fluorescence from the surface of a microsphere due to the cleavage of an attached fluorescent protease substrate by a suitable protease enzyme. The assay format can be adapted to any site or protein-specific protease of interest and results can be measured in both real time and as endpoint fluorescence assays on a flow cytometer. Endpoint assays are easily adapted to microplate format for flow cytometry high-throughput analysis and inhibitor screening.

Topics: Animals; Biotinylation; Flow Cytometry; Fluorescence Resonance Energy Transfer; Green Fluorescent Proteins; High-Throughput Screening Assays; Humans; Inflammation; Kinetics; Microspheres; Peptide Hydrolases; Peptides; Reproducibility of Results; Temperature

Four coumarins were isolated from the EtOAc extract of the flower-tops of Santolina oblongifolia Boiss. (Compositae). They were identified as 7-methoxycoumarin (herniarin) (1), 6,7-dihydroxycoumarin (aesculetin) (2), 6-methoxy-7-glucosidylcoumarin (scopolin) (3), and 6-hydroxy-7-methoxycoumarin (scopoletin) (4). This is the first report of the isolation of aesculetin and scopolin from the genus Santolina. The isolated coumarins showed marked activity as inhibitors of eicosanoid-release from ionophore-stimulated mouse peritoneal macrophages.

Topics: Animals; Anti-Inflammatory Agents; Calcimycin; Carrageenan; Chromatography, Thin Layer; Coumarins; Dinoprostone; Inflammation; Ionophores; Leukotrienes; Macrophages, Peritoneal; Magnetic Resonance Spectroscopy; Mice; Plant Extracts; Plants, Medicinal; Rats; Rats, Wistar; Spain; Spectrophotometry, Ultraviolet