scutellarein and chrysin

Baicalein and scutellarein are bioactive flavones found in the medicinal plant Scutellaria baicalensis Georgi, used in traditional Chinese medicine. Extensive previous work has demonstrated the broad biological activity of these flavonoids, such as antifibrotic, antiviral and anticancer properties. However, their supply from plant material is insufficient to meet demand. Here, to provide an alternative production source and increase production levels of these flavones, we engineered an artificial pathway in an Escherichia coli cell factory for the first time. By first reconstructing the plant flavonoid biosynthetic pathway genes from five different species: phenylalanine ammonia lyase from Rhodotorula toruloides (PAL), 4-coumarate-coenzyme A ligase from Petroselinum crispum (4CL), chalcone synthase from Petunia hybrida (CHS), chalcone isomerase from Medicago sativa (CHI) and an oxidoreductase flavone synthase I from P. crispum (FNSI), production of the intermediates chrysin and apigenin was achieved by feeding phenylalanine and tyrosine as precursors. By comparative analysis of various versions of P450s, a construction expressing 2B1 incorporated with a 22-aa N-terminal truncated flavone C-6 hydroxylase from S. baicalensis (F6H) and partner P450 reductase from Arabidopsis thaliana (AtCPR) was found most effective for production of both baicalein (8.5 mg/L) and scutellarein (47.1 mg/L) upon supplementation with 0.5 g/L phenylalanine and tyrosine in 48 h of fermentation. Finally, optimization of malonyl-CoA availability further increased the production of baicalein to 23.6 mg/L and scutellarein to 106.5 mg/L in a flask culture. This report presents a significant advancement of flavone synthetic production and provides foundation for production of other flavones in microbial hosts.

Topics: Apigenin; Biosynthetic Pathways; Escherichia coli; Flavanones; Flavonoids; Malonyl Coenzyme A; Metabolic Engineering; Phenylalanine; Plants; Scutellaria baicalensis; Tyrosine

Topics: Andrographis; Antineoplastic Agents, Phytogenic; Apigenin; Apocynaceae; Asia, Southeastern; Biflavonoids; Bignoniaceae; Catechin; Diterpenes; Ethnopharmacology; Flavanones; Flavonoids; Hep G2 Cells; Humans; Lamiaceae; Liver Neoplasms; Metabolomics; Plants, Medicinal; Proanthocyanidins

Flavone glucuronide isomers of five flavones (chrysin, apigenin, luteolin, baicalein, and scutellarein) were differentiated by collision-induced dissociation of [Co(II) (flavone-H) (4,7-diphenyl-1,10-phenanthroline)(2)](+) complexes. The complexes were generated via postcolumn addition of a metal-ligand solution after separation of the glucuronide products generated upon incubation of each flavone with an array of uridine 5'-diphosphate (UDP)-glucuronosyltransferase (UGT) isozymes. Elucidation of the glucuronide isomers allowed a systematic investigation of the regioselectivity of 12 human UGT isozymes, including 8 UGT1A and 4 UGT2B isozymes. Glucuronidation of the 7-OH position was the preferred site for all the flavones except for luteolin, which possessed adjacent hydroxyl groups on the B ring. For all flavones and UGT isozymes, glucuronidation of the 5-OH position was never observed. As confirmed by the metal complexation/MS/MS strategy, glucuronidation of the 6-OH position only occurred for baicalein and scutellarein when incubated with three of the UGT isozymes.

Topics: Apigenin; Biotransformation; Chromatography, High Pressure Liquid; Coordination Complexes; Flavanones; Flavonoids; Glucuronides; Glucuronosyltransferase; Isoenzymes; Isomerism; Luteolin; Tandem Mass Spectrometry

Up to now, the metabolism of hispidulin (5,7,4'-trihydroxy-6-methoxyflavone), a potent ligand of the central human benzodiazepine receptor, has not been investigated. To elucidate the metabolism of hispidulin in the large intestine, its biotransformation by the pig caecal microflora was studied. In addition, the efficiency of the pig caecal microflora to degrade galangin (3,5,7-trihydroxyflavone), kaempferol (3,5,7,4'-tetrahydroxyflavone), apigenin (5,7,4'-trihydroxyflavone), and luteolin (5,7,3',4'-tetrahydroxyflavone) was investigated. Identification of the formed metabolites was performed by high-performance liquid chromatography (HPLC)-diode array detection, HPLC-electrospray ionization-tandem mass spectrometry, and high-resolution gas chromatography-mass spectrometry. The caecal microflora transformed hispidulin to scutellarein (5,6,7,4'-tetrahydroxyflavone), an effective alpha-glucosidase inhibitor, and 3-(4-hydroxyphenyl)-propionic acid; galangin to phenylacetic acid and phloroglucinol; kaempferol to 4-hydroxyphenylacetic acid, phloroglucinol, and 4-methylphenol; apigenin to 3-(4-hydroxyphenyl)-propionic acid and 3-phenylpropionic acid, and luteolin to 3-(3-hydroxyphenyl)-propionic acid, respectively. To elucidate to what extent different hydroxylation patterns on the B-ring influence the degradation degree of flavonoids, the conversions of galangin and kaempferol as well as that of apigenin and luteolin were compared with those of quercetin (3,5,7,3',4'-pentahydroxyflavone) and chrysin (5,7-dihydroxyflavone), respectively. Regardless of the flavonoid subclass, the presence of a hydroxy group at the 4'-position seems to be a prerequisite for fast breakdown. An additional hydroxy group at the B-ring did not affect the degradation degree.

Topics: Animals; Apigenin; Bacteria; Cecum; Chromatography, High Pressure Liquid; Flavones; Flavonoids; Gas Chromatography-Mass Spectrometry; Humans; Kaempferols; Kinetics; Luteolin; Models, Animal; Quercetin; Spectrometry, Mass, Electrospray Ionization; Swine

Twenty-six flavonoids were isolated from Scutellaria baicalensis. Their affinities for the benzodiazepine (BDZ) binding site of GABA A receptor have been studied using [ 3H]flunitrazepam binding to rat cortical membranes in vitro. The structure-activity relationships suggested that 2'-OH flavones exhibited the most potent binding affinity, which could lead to the design and discovery of new BDZ receptor ligands.

Topics: Animals; Apigenin; Binding Sites; Flavanones; Flavonoids; Flunitrazepam; GABA Modulators; Glucuronates; Molecular Structure; Plant Extracts; Quantitative Structure-Activity Relationship; Rats; Rats, Sprague-Dawley; Receptors, GABA-A; Scutellaria baicalensis